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Sample records for fiberoptic dna probe

  1. A new method of preparing fiber-optic DNA biosensor and its array for gene detection

    Institute of Scientific and Technical Information of China (English)

    2001-01-01

    A new method of preparing fiber-optic DNA biosensor and its arrayfor the simultaneous detection of multiple genes is described. The optical fibers were first treated with poly-l-lysine, and then were made into fiber-optic DNA biosensors by adsorbing and immobilizing the oligonucleotide probe on its end. By assembling the fiber-optic DNA biosensors in a bundle in which each fiber carried a different DNA probe, the fiber-optic DNA biosensor array was well prepared. Hybridization of fluorescent- labeled cDNA of p53 gene, N-ras gene and Rb1 gene to the DNA array was monitored by CCD camera. A good result was achieved.

  2. Ultrathin lensed fiber-optic probe for optical coherence tomography.

    Science.gov (United States)

    Qiu, Y; Wang, Y; Belfield, K D; Liu, X

    2016-06-01

    We investigated and validated a novel method to develop ultrathin lensed fiber-optic (LFO) probes for optical coherence tomography (OCT) imaging. We made the LFO probe by attaching a segment of no core fiber (NCF) to the distal end of a single mode fiber (SMF) and generating a curved surface at the tip of the NCF using the electric arc of a fusion splicer. The novel fabrication approach enabled us to control the length of the NCF and the radius of the fiber lens independently. By strategically choosing these two parameters, the LFO probe could achieve a broad range of working distance and depth of focus for different OCT applications. A probe with 125μm diameter and lateral resolution up to 10μm was demonstrated. The low-cost, disposable and robust LFO probe is expected to have great potential for interstitial OCT imaging.

  3. Hollow-core photonic crystal fiber-optic probes for Raman spectroscopy.

    Science.gov (United States)

    Konorov, Stanislav O; Addison, Christopher J; Schulze, H Georg; Turner, Robin F B; Blades, Michael W

    2006-06-15

    We have implemented a new Raman fiber-optic probe design based on a hollow-core photonic-crystal excitation fiber surrounded by silica-core collection fibers. The photonic-crystal fiber offers low attenuation at the pump radiation wavelength, mechanical flexibility, high radiation stability, and low background noise. Because the excitation beam is transmitted through air inside the hollow-core fiber, silica Raman scattering is much reduced, improving the quality of the spectra obtained using probes of this design. Preliminary results show that the new probe design decreases the Raman background from the silica by approximately an order of magnitude compared to solid-core silica Raman probes.

  4. Probing the ultimate limit of fiber-optic strain sensing.

    Science.gov (United States)

    Gagliardi, G; Salza, M; Avino, S; Ferraro, P; De Natale, P

    2010-11-19

    The measurement of relative displacements and deformations is important in many fields such as structural engineering, aerospace, geophysics, and nanotechnology. Optical-fiber sensors have become key tools for strain measurements, with sensitivity limits ranging between 10(-9) and 10(-6)ε hertz (Hz)(-1/2) (where ε is the fractional length change). We report on strain measurements at the 10(-13)ε-Hz(-1/2) level using a fiber Bragg-grating resonator with a diode-laser source that is stabilized against a quartz-disciplined optical frequency comb, thus approaching detection limits set by thermodynamic phase fluctuations in the fiber. This scheme may provide a route to a new generation of strain sensors that is entirely based on fiber-optic systems, which are aimed at measuring fundamental physical quantities; for example, in gyroscopes, accelerometers, and gravity experiments.

  5. Interferometric measurement of injection nozzles using ultra-small fiber-optical probes

    Institute of Scientific and Technical Information of China (English)

    Tilo Pfeifer; Robert Schmitt; Niels K(o)nig; Guilherme Francisco Mallmann

    2011-01-01

    The measurement of boreholes with diameters smaller than 500 pm is a demanding task that cannot be performed using state-of-the-art production metrology.In this letter,a miniaturized fiber probe with a diameter of 80 pm is presented.A probe is used for low-coherence interferometry to conduct highly precise measurements of form deviations of small boreholes.Measurements conducted in nozzles are also presented.The results prove the potential of the fiber-optical sensor for quality inspection of high-precision parts,such as injection nozzles,for common-rail diesel engines.

  6. A fiber-optic probe for particle sizing in concentrated suspensions

    Science.gov (United States)

    Dhadwal, Harbans S.; Ansari, Rafat R.; Meyer, William V.

    1991-01-01

    A fiber-optic probe employing two monomode optical fibers, one for transmitting a Gaussian laser beam to the scattering volume and the second, positioned at some backscatter angle, for receiving the scattered light is described. Performance and suitability of the system for a process control environment is assessed by studying a suspension of polystyrene latex particles over a wide range of sizes and concentrations. The results show that the probe is ideal for a process control environment in industrial and laboratory applications. Particle size is recovered, without any additional corrections for multiple light scattering, in concentrations containing up to 10 percent solids of 39-nm polystyrene latex spheres.

  7. Fiber-optic laser Doppler turbine tip clearance probe.

    Science.gov (United States)

    Büttner, Lars; Pfister, Thorsten; Czarske, Jürgen

    2006-05-01

    A laser Doppler based method for in situ single blade tip clearance measurements of turbomachines with high precision is presented for what we believe is the first time. The sensor is based on two superposed fanlike interference fringe systems generated by two laser wavelengths from a fiber-coupled, passive, and therefore compact measurement head employing diffractive optics. Tip clearance measurements at a transonic centrifugal compressor performed during operation at 50,000 rpm (833 Hz, 586 m/s tip speed) are reported. At these speeds the measured uncertainty of the tip position was less than 20 microm, a factor of 2 more accurate than that of capacitive probes. The sensor offers great potential for in situ and online high-precision tip clearance measurements of metallic and nonmetallic turbine blades.

  8. Interferometric strain measurements with a fiber-optic probe

    Science.gov (United States)

    Burnham-Fay, E. D.; Jacobs-Perkins, D. W.; Ellis, J. D.

    2015-09-01

    Experience at the Laboratory for Laser Energetics has shown that broadband base vibrations make it difficult to position cryogenic inertial confinement fusion targets. These effects must be mitigated for National Ignition Facility-scale targets; to this end an active vibration stabilization system is proposed. A single-mode optical fiber strain probe and a novel fiber contained heterodyne interferometer have been developed as a position feedback sensor for the vibration control system. A resolution limit of 54.5 nƐ; is measured with the optical strain gauge, limited by the lock-in amplifier. Experimental measurements of the sensor that show good agreement with reference resistive strain gauge measurements are presented.

  9. DNA probe for lactobacillus delbrueckii

    Energy Technology Data Exchange (ETDEWEB)

    Delley, M.; Mollet, B.; Hottinger, H. (Nestle Research Centre, Lausanne (Switzerland))

    1990-06-01

    From a genomic DNA library of Lactobacillus delbrueckii subsp. bulgaricus, a clone was isolated which complements a leucine auxotrophy of an Escherichia coli strain (GE891). Subsequent analysis of the clone indicated that it could serve as a specific DNA probe. Dot-blot hybridizations with over 40 different Lactobacillus strains showed that this clone specifically recognized L. delbrueckii subsp. delbrueckii, bulgaricus, and lactis. The sensitivity of the method was tested by using an {alpha}-{sup 32}P-labeled probe.

  10. Novel DNA probes for sensitive DNA detection

    OpenAIRE

    Richardson, James Alistair

    2010-01-01

    The ability to detect and interrogate DNA sequences allows further understanding and\\ud diagnosis of genetic disease. The ability to perform such analysis of genetic material\\ud requires highly selective and reliable technologies. Furthermore techniques which can use\\ud simple and cheap equipment allow the use of such technologies for point of care analysis.\\ud \\ud Described in this thesis are two novel DNA probe systems designed for mutation\\ud discrimination and sequence recognition of PCR ...

  11. Fiber-Optic Imaging Probe Developed for Space Used to Detect Diabetes Through the Eye

    Science.gov (United States)

    Ansari, Rafat R.; Chenault, Michelle V.; Datiles, Manuel B., III; Sebag, J.; Suh, Kwang I.

    2000-01-01

    Approximately 16 million Americans have diabetes mellitus, which can severely impair eyesight by causing cataracts, diabetic retinopathy, and glaucoma. Cataracts are 1.6 times more common in people with diabetes than in those without diabetes, and cataract extraction is the only surgical treatment. In many cases, diabetes-related ocular pathologies go undiagnosed until visual function is compromised. This ongoing pilot project seeks to study the progression of diabetes in a unique animal model by monitoring changes in the lens with a safe, sensitive, dynamic light-scattering probe. Dynamic light scattering (DLS), has the potential to diagnose cataracts at the molecular level. Recently, a new DLS fiber-optic probe was developed at the NASA Glenn Research Center at Lewis Field for noncontact, accurate, and extremely sensitive particle-sizing measurements in fluid dispersions and suspensions (ref. 1). This compact, portable, and rugged probe is free of optical alignment, offers point-and-shoot operation for various online field applications and challenging environments, and yet is extremely flexible in regards to sample container sizes, materials, and shapes. No external vibration isolation and no index matching are required. It can measure particles as small as 1 nm and as large as few micrometers in a wide concentration range from very dilute (waterlike) dispersions to very turbid (milklike) suspensions. It is safe and fast to use, since it only requires very low laser power (10 nW to 3 mW) with very short data acquisition times (2 to 10 sec).

  12. DETECTION OF DNA DAMAGE USING A FIBEROPTIC BIOSENSOR

    Science.gov (United States)

    A rapid and sensitive fiber optic biosensor assay for radiation-induced DNA damage is reported. For this assay, a biotin-labeled capture oligonucleotide (38 mer) was immobilized to an avidin-coated quartz fiber. Hybridization of a dye-labeled complementary sequence was observed...

  13. Fiber-optical sensor with miniaturized probe head and nanometer accuracy based on spatially modulated low-coherence interferogram analysis.

    Science.gov (United States)

    Depiereux, Frank; Lehmann, Peter; Pfeifer, Tilo; Schmitt, Robert

    2007-06-10

    Fiber-optical sensors have some crucial advantages compared with rigid optical systems. They allow miniaturization and flexibility of system setups. Nevertheless, optical principles such as low-coherence interferometry can be realized by use of fiber optics. We developed and realized an approach for a fiber-optical sensor, which is based on the analysis of spatially modulated low-coherence interferograms. The system presented consists of three units, a miniaturized sensing probe, a broadband fiber-coupled light source, and an adapted Michelson interferometer, which is used as an optical receiver. Furthermore, the signal processing procedure, which was developed for the interferogram analysis in order to achieve nanometer measurement accuracy, is discussed. A system prototype has been validated thoroughly in different experiments. The results approve the accuracy of the sensor.

  14. Design and characterization of a novel multimodal fiber-optic probe and spectroscopy system for skin cancer applications

    Science.gov (United States)

    Sharma, Manu; Marple, Eric; Reichenberg, Jason; Tunnell, James W.

    2014-08-01

    The design and characterization of an instrument combining Raman, fluorescence, and reflectance spectroscopic modalities is presented. Instrument development has targeted skin cancer applications as a novel fiber-optic probe has been specially designed to interrogate cutaneous lesions. The instrument is modular and both its software and hardware components are described in depth. Characterization of the fiber-optic probe is also presented, which details the probe's ability to measure diagnostically important parameters such as intrinsic fluorescence and absorption and reduced scattering coefficients along with critical performance metrics such as high Raman signal-to-noise ratios at clinically practical exposure times. Validation results using liquid phantoms show that the probe and system can extract absorption and scattering coefficients with less than 10% error. As the goal is to use the instrument for the clinical early detection of skin cancer, preliminary clinical data are also presented, which indicates our system's ability to measure physiological quantities such as relative collagen and nicotinamide adenine dinucleotide concentration, oxygen saturation, blood volume fraction, and mean vessel diameter.

  15. Probe and method for DNA detection

    Science.gov (United States)

    Yeh, Hsin-Chih; Werner, James Henry; Sharma, Jaswinder Kumar; Martinez, Jennifer Suzanne

    2013-07-02

    A hybridization probe containing two linear strands of DNA lights up upon hybridization to a target DNA using silver nanoclusters that have been templated onto one of the DNA strands. Hybridization induces proximity between the nanoclusters on one strand and an overhang on the other strand, which results in enhanced fluorescence emission from the nanoclusters.

  16. Effect of low intensity laser interaction with human skin fibroblast cells using fiber-optic nano-probes.

    Science.gov (United States)

    Pal, Gopalendu; Dutta, Ashim; Mitra, Kunal; Grace, Michael S; Amat, Albert; Romanczyk, Tara B; Wu, Xingjia; Chakrabarti, Kristi; Anders, Juanita; Gorman, Erik; Waynant, Ronald W; Tata, Darrell B

    2007-03-01

    Over the past forty years, many efforts have been devoted to study low power laser light interactions with biological systems. Some of the investigations were performed in-vitro, on bulk cell populations. Our present work was undertaken to apply specially engineered fiber-optic based nano-probes for the precise delivery of laser light on to a single cell and to observe production of low power laser light induced reactive oxygen species (ROS). A normal human skin fibroblast (NHF) cell line was utilized in this investigation and the cells were irradiated under two different schemes of exposure: (1) an entire NHF cell population within a Petri dish using a fan beam methodology, and (2) through the precise delivery of laser energy on to a single NHF cell using fiber-optic nano-probe. Photobiostimulative studies were conducted through variation of laser intensity, exposure time, and the energy dose of exposure. Laser irradiation induced enhancement in the rate of cell proliferation was observed to be dependent on laser exposure parameters and the method of laser delivery. The total energy dose (fluence) had a greater influence on the enhancement in the rate of cellular proliferation than compared to laser intensity. The enhancement in the growth rate was observed to have a finite life-time of several days after the initial laser exposure. Fluorescent life-time imaging of ROS was performed during the nano-based single cell exposure method. The kinetics of ROS generation was found to depend strongly on the laser fluence and not on the laser intensity.

  17. Development and application of DNA molecular probes

    Directory of Open Access Journals (Sweden)

    Priya Vizzini

    2017-02-01

    Full Text Available The development of DNA probes started from 1950's for diagnostic purposes and it is still growing. DNA probes are applied in several fields such as food, medical, veterinary, environment and security, with the aim of prevention, diagnosis and treatment. The use of DNA probes permits microorganism identification, including pathogen detection, and their quantification when used in specific systems. Various techniques obtained success by the utilization of specific DNA probes, that allowed the obtainment of rapid and specific results. From PCR, qPCR and blotting techniques that were first used in well equipped laboratories to biosensors such as fiber optic, surface plasmon resonance (SPR, electrochemical, and quartz crystal microbalance (QCM biosensors that use different transduction systems. This review describes i the design and production of primers and probes, and their utilization from the traditional techniques to the new emerging techniques like biosensors used in current applications; ii the possibility to use labelled-free probes and probes labelled with an enzyme/fluorophore, etc.; iii the different sensitivity obtained by using specific systems; and iv the advantage obtained by using biosensors.

  18. Development of DNA probes for Candida albicans

    Energy Technology Data Exchange (ETDEWEB)

    Cheung, L.L.; Hudson, J.B.

    1988-07-01

    An attempt was made to produce DNA probes that could be used as a rapid and efficient means of detecting candidiasis (invasive Candida infection) in immunocompromised patients. Whole DNA from Candida albicans was digested with restriction endonuclease, and the resulting fragments were randomly cloned into a plasmid vector. Several recombinant plasmids were evaluated for cross-hybridization to various other Candida species, other fungal DNAs, and to nonfungal DNAs. Cross reactions were observed between the probes and different yeasts, but none with unrelated DNAs. Some recombinants were genus-specific, and two of these were applied to the analysis of C. albicans growth curves. It became evident that, although both /sup 32/P- and biotin-labelled probes could be made quite sensitive, a possible limitation in their diagnostic potential was the poor liberation of Candida DNA from cells. Thus, better methods of treatment of clinical specimens will be required before such probes will be useful in routine diagnosis.

  19. DNA binding hydroxyl radical probes

    Energy Technology Data Exchange (ETDEWEB)

    Tang, Vicky J.; Konigsfeld, Katie M.; Aguilera, Joe A. [Department of Radiology, University of California at San Diego, 9500 Gilman Drive, La Jolla, CA 92093-0610 (United States); Milligan, Jamie R., E-mail: jmilligan@ucsd.edu [Department of Radiology, University of California at San Diego, 9500 Gilman Drive, La Jolla, CA 92093-0610 (United States)

    2012-01-15

    The hydroxyl radical is the primary mediator of DNA damage by the indirect effect of ionizing radiation. It is a powerful oxidizing agent produced by the radiolysis of water and is responsible for a significant fraction of the DNA damage associated with ionizing radiation. There is therefore an interest in the development of sensitive assays for its detection. The hydroxylation of aromatic groups to produce fluorescent products has been used for this purpose. We have examined four different chromophores, which produce fluorescent products when hydroxylated. Of these, the coumarin system suffers from the fewest disadvantages. We have therefore examined its behavior when linked to a cationic peptide ligand designed to bind strongly to DNA. - Highlights: > Examined four aromatic groups as a means to detect hydroxyl radicals by fluorescence. > Coumarin system suffers from the fewest disadvantages. > Characterized its reactivity when linked to a hexa-arginine peptide.

  20. Chromosome-specific DNA Repeat Probes

    Energy Technology Data Exchange (ETDEWEB)

    Baumgartner, Adolf; Weier, Jingly Fung; Weier, Heinz-Ulrich G.

    2006-03-16

    In research as well as in clinical applications, fluorescence in situ hybridization (FISH) has gained increasing popularity as a highly sensitive technique to study cytogenetic changes. Today, hundreds of commercially available DNA probes serve the basic needs of the biomedical research community. Widespread applications, however, are often limited by the lack of appropriately labeled, specific nucleic acid probes. We describe two approaches for an expeditious preparation of chromosome-specific DNAs and the subsequent probe labeling with reporter molecules of choice. The described techniques allow the preparation of highly specific DNA repeat probes suitable for enumeration of chromosomes in interphase cell nuclei or tissue sections. In addition, there is no need for chromosome enrichment by flow cytometry and sorting or molecular cloning. Our PCR-based method uses either bacterial artificial chromosomes or human genomic DNA as templates with {alpha}-satellite-specific primers. Here we demonstrate the production of fluorochrome-labeled DNA repeat probes specific for human chromosomes 17 and 18 in just a few days without the need for highly specialized equipment and without the limitation to only a few fluorochrome labels.

  1. Study of polarization properties of fiber-optics probes with use of a binary phase plate.

    Science.gov (United States)

    Alferov, S V; Khonina, S N; Karpeev, S V

    2014-04-01

    We conduct a theoretical and experimental study of the distribution of the electric field components in the sharp focal domain when rotating a zone plate with a π-phase jump placed in the focused beam. Comparing the theoretical and experimental results for several kinds of near-field probes, an analysis of the polarization sensitivity of different types of metal-coated aperture probes is conducted. It is demonstrated that with increasing diameter of the non-metal-coated tip part there occurs an essential redistribution of sensitivity in favor of the transverse electric field components and an increase of the probe's energy throughput.

  2. Universal microbial diagnostics using random DNA probes

    Science.gov (United States)

    Aghazadeh, Amirali; Lin, Adam Y.; Sheikh, Mona A.; Chen, Allen L.; Atkins, Lisa M.; Johnson, Coreen L.; Petrosino, Joseph F.; Drezek, Rebekah A.; Baraniuk, Richard G.

    2016-01-01

    Early identification of pathogens is essential for limiting development of therapy-resistant pathogens and mitigating infectious disease outbreaks. Most bacterial detection schemes use target-specific probes to differentiate pathogen species, creating time and cost inefficiencies in identifying newly discovered organisms. We present a novel universal microbial diagnostics (UMD) platform to screen for microbial organisms in an infectious sample, using a small number of random DNA probes that are agnostic to the target DNA sequences. Our platform leverages the theory of sparse signal recovery (compressive sensing) to identify the composition of a microbial sample that potentially contains novel or mutant species. We validated the UMD platform in vitro using five random probes to recover 11 pathogenic bacteria. We further demonstrated in silico that UMD can be generalized to screen for common human pathogens in different taxonomy levels. UMD’s unorthodox sensing approach opens the door to more efficient and universal molecular diagnostics. PMID:27704040

  3. Fiber-optic confocal microscopy using a miniaturized needle-compatible imaging probe

    Science.gov (United States)

    Pillai, Rajesh S.; Lorenser, Dirk; Sampson, David D.

    2011-05-01

    We report on the design and implementation of a 350 μm-diameter confocal imaging probe based on gradient-index (GRIN) optics and a fiber-based scanning arrangement. The form factor of the probe is such that it can potentially be inserted into a 22-gauge hypodermic needle to perform high-resolution confocal fluorescence imaging in solid tissues. We introduce a simple scanning arrangement based on lensed fiber, which eliminates off-axis aberrations induced by conventional scanning optics and is suitable for integration into a compact hand-held unit. We present the details of the optical design and experimental verification of the performance of the optical system. The measured lateral resolution of ~700 nm is in agreement with the optical design and is the highest resolution reported for a confocal fluorescence imaging probe of this size. Further, we demonstrate the imaging capability of the probe by obtaining high-resolution images of fluorescently labeled muscle fibers.

  4. Development of Fiber-Optic Humidity Sensor Probe with Gelatin Cladding

    Directory of Open Access Journals (Sweden)

    Akhiruddin Maddu

    2010-10-01

    Full Text Available Humidity sensor based on optical fiber with gelatin cladding has been developed. In this humidity sensor probe, the origin cladding of optical fiber is replaced by gelatin coating as humidity sensitive cladding. Testing of the optical fiber sensor probe was conducted by measuring of light intensity transmitted on the optical fiber probe for each variation of different humidity treatments. Response of the optical fiber sensor probe measured from 42%RH to 99%RH, the results show an optical transmission curve varied with relative humidity (RH. Optical transmission in the optical fiber probe increase with RH value at a specific wavelength range, that is from green to red spectrum bands (500 nm - 700 nm, where a significant variation from 600 nm to 650 nm in yellow to red spectrum bands. Wavelength where is a maximum intensity of optical transmission occurs at 610 nm. Therefore, the optical fiber humidity sensor probe could response humidity form 42%RH to 99%RH with the best response in humidity range of 60%RH to 72%RH that is have a good  linearity and sensitivity

  5. Fiber-optic probe for noninvasive real-time determination of tissue optical properties at multiple wavelengths.

    Science.gov (United States)

    Dam, J S; Pedersen, C B; Dalgaard, T; Fabricius, P E; Aruna, P; Andersson-Engels, S

    2001-03-01

    We present a compact, fast, and versatile fiber-optic probe system for real-time determination of tissue optical properties from spatially resolved continuous-wave diffuse reflectance measurements. The system collects one set of reflectance data from six source-detector distances at four arbitrary wavelengths with a maximum overall sampling rate of 100 Hz. Multivariate calibration techniques based on two-dimensional polynomial fitting are employed to extract and display the absorption and reduced scattering coefficients in real-time mode. The four wavelengths of the current configuration are 660, 785, 805, and 974 nm, respectively. Cross-validation tests on a 6 x 7 calibration matrix of Intralipid-dye phantoms showed that the mean prediction error at, e.g., 785 nm was 2.8% for the absorption coefficient and 1.3% for the reduced scattering coefficient. The errors are relative to the range of the optical properties of the phantoms at 785 nm, which were 0-0.3/cm for the absorption coefficient and 6-16/cm for the reduced scattering coefficient. Finally, we also present and discuss results from preliminary skin tissue measurements.

  6. Novel fiber-optical interferometer with miniaturized probe for in-hole measurements

    Institute of Scientific and Technical Information of China (English)

    Robed Schmitt; Tilo Pfeifer; Frank Depiereux; Niels K(o)nig

    2008-01-01

    Today,micro-system technology and the development of new MEMS(Micro-Eleetro-Mechanical Systems)ate emerging rapidly.In order for this development to become a SUCCESS in the long run,measurement systems have to ensure product quality.Most often,MEMS have to be tested by means of functionality or destructive tests.One reason for this is that there are no suitable systems or sensing probes available which can be used for the measurement of quasi inaccessible features like small holes or cavities.We present a measurement system that could be used for these kinds of measurements.The system combines a fiber optical,miniaturized sensing probe with low-coherence intcrferometry,so that absolute distance measurements with nanometcr accuracy are possible.

  7. Human Oral Mucosa Tissue-Engineered Constructs Monitored by Raman Fiber-Optic Probe

    OpenAIRE

    Khmaladze, Alexander; Kuo, Shiuhyang; Kim, Roderick Y; Matthews, Robert V.; Marcelo, Cynthia L.; Feinberg, Stephen E.; Morris, Michael D.

    2014-01-01

    In maxillofacial and oral surgery, there is a need for the development of tissue-engineered constructs. They are used for reconstructions due to trauma, dental implants, congenital defects, or oral cancer. A noninvasive monitoring of the fabrication of tissue-engineered constructs at the production and implantation stages done in real time is extremely important for predicting the success of tissue-engineered grafts. We demonstrated a Raman spectroscopic probe system, its design and applicati...

  8. Quantitative optical coherence elastography based on fiber-optic probe with integrated Fabry-Perot force sensor

    Science.gov (United States)

    Qiu, Yi; Wang, Yahui; Xu, Yiqing; Chandra, Namas; Haorah, James; Hubbi, Basil; Pfister, Bryan J.; Liu, Xuan

    2016-03-01

    Optical coherence tomography (OCT) is a versatile imaging technique and has great potential in tissue characterization for breast cancer diagnosis and surgical guidance. In addition to structural difference, cancerous breast tissue is usually stiffer compared to normal adipose breast tissue. However, previous studies on compression optical coherence elastography (OCE) are qualitative rather than quantitative. It is challenging to identify the cancerous status of tissue based on qualitative OCE results obtained from different measurement sessions or from different patients. Therefore, it is critical to develop technique that integrates structural imaging and force sensing, for quantitative elasticity characterization of breast tissue. In this work, we demonstrate a quantitative OCE (qOCE) microsurgery device which simultaneously quantifies force exerted to tissue and measures the resultant tissue deformation. The qOCE system is based on a spectral domain OCT engine operated at 1300 nm and a probe with an integrated Febry-Perot (FP) interferometric cavity at its distal end. The FP cavity is formed by the cleaved end of the lead-in fiber and the end surface of a GRIN lens which allows light to incident into tissue for structural imaging. The force exerted to tissue is quantified by the change of FP cavity length which is interrogated by a fiber-optic common-paths phase resolved OCT system with sub-nanometer sensitivity. Simultaneously, image of the tissue structure is acquired from photons returned from tissue through the GRIN lens. Tissue deformation is obtained through Doppler analysis. Tissue elasticity can be quantified by comparing the force exerted and tissue deformation.

  9. Fiber-optic polarization diversity detection for rotary probe optical coherence tomography.

    Science.gov (United States)

    Lee, Anthony M D; Pahlevaninezhad, Hamid; Yang, Victor X D; Lam, Stephen; MacAulay, Calum; Lane, Pierre

    2014-06-15

    We report a polarization diversity detection scheme for optical coherence tomography with a new, custom, miniaturized fiber coupler with single mode (SM) fiber inputs and polarization maintaining (PM) fiber outputs. The SM fiber inputs obviate matching the optical lengths of the X and Y OCT polarization channels prior to interference and the PM fiber outputs ensure defined X and Y axes after interference. Advantages for this scheme include easier alignment, lower cost, and easier miniaturization compared to designs with free-space bulk optical components. We demonstrate the utility of the detection system to mitigate the effects of rapidly changing polarization states when imaging with rotating fiber optic probes in Intralipid suspension and during in vivo imaging of human airways.

  10. Fiber optic chemical sensors: The evolution of high- density fiber-optic DNA microarrays

    Science.gov (United States)

    Ferguson, Jane A.

    2001-06-01

    and O2. This sensor is useful for monitoring bioprocesses such as (beer) fermentation and for clinical situations such as blood gas analysis. DNA sensors were created by attaching short single strands of DNA (probes) to the fiber tip. A matching single strand (target) forms a strong interacting pair with the probe upon contact. The target strands in a sample are labeled with a fluorescent dye. When a probe-target pair is formed and excitation light is sent down the fiber, the fiber bearing the pair emits light that is captured and detected. A high density DNA array was created by isolating thousands of discrete DNA sensors on the tip of an imaging optical fiber. This array was made possible by the formation of microwells on the imaging fiber tip. Microspheres functionalized with DNA were placed in the wells of the fiber and each microsphere was independently and simultaneously monitored. (Abstract shortened by UMI.)

  11. Raman fiberoptic probe for monitoring human tissue engineered oral mucosa constructs

    Science.gov (United States)

    Khmaladze, Alexander; Kuo, Shiuhyang; Okagbare, Paul; Marcelo, Cynthia L.; Feinberg, Stephen E.; Morris, Michael D.

    2013-02-01

    In oral and maxillofacial surgery, there is a need for tissue engineered constructs for dental implants, reconstructions due to trauma, oral cancer or congenital defects. A non-invasive quality monitoring of the fabrication of tissue engineered constructs during their production and implantation is a required component of any successful tissue engineering technique. We demonstrate the design and application of a Raman spectroscopic probe for rapid and noninvasive monitoring of Ex Vivo Produced Oral Mucosa Equivalent constructs (EVPOMEs). We conducted in vivo studies to identify Raman spectroscopic failure indicators for EVPOMEs (already developed in vitro), and found that Raman spectra of EVPOMEs exposed to thermal stress showed correlation of the band height ratio of CH2 deformation to phenylalanine ring breathing modes, providing a Raman metric to distinguish between viable and nonviable constructs. This is the first step towards the ultimate goal to design a stand-alone system, which will be usable in a clinical setting, as the data processing and analysis will be performed with minimal user intervention, based on already established and tested Raman spectroscopic indicators for EVPOMEs.

  12. Human oral mucosa tissue-engineered constructs monitored by Raman fiber-optic probe.

    Science.gov (United States)

    Khmaladze, Alexander; Kuo, Shiuhyang; Kim, Roderick Y; Matthews, Robert V; Marcelo, Cynthia L; Feinberg, Stephen E; Morris, Michael D

    2015-01-01

    In maxillofacial and oral surgery, there is a need for the development of tissue-engineered constructs. They are used for reconstructions due to trauma, dental implants, congenital defects, or oral cancer. A noninvasive monitoring of the fabrication of tissue-engineered constructs at the production and implantation stages done in real time is extremely important for predicting the success of tissue-engineered grafts. We demonstrated a Raman spectroscopic probe system, its design and application, for real-time ex vivo produced oral mucosa equivalent (EVPOME) constructs noninvasive monitoring. We performed in vivo studies to find Raman spectroscopic indicators for postimplanted EVPOME failure and determined that Raman spectra of EVPOMEs preexposed to thermal stress during manufacturing procedures displayed correlation of the band height ratio of CH2 deformation to phenylalanine ring breathing modes, giving a Raman metric to distinguish between healthy and compromised postimplanted constructs. This study is the step toward our ultimate goal to develop a stand-alone system, to be used in a clinical setting, where the data collection and analysis are conducted on the basis of these spectroscopic indicators with minimal user intervention.

  13. Fiber-optic system for dual-modality imaging of glucose probes 18F-FDG and 6-NBDG in atherosclerotic plaques.

    Directory of Open Access Journals (Sweden)

    Raiyan T Zaman

    Full Text Available Atherosclerosis is a progressive inflammatory condition that underlies coronary artery disease (CAD-the leading cause of death in the United States. Thus, the ultimate goal of this research is to advance our understanding of human CAD by improving the characterization of metabolically active vulnerable plaques within the coronary arteries using a novel catheter-based imaging system. The aims of this study include (1 developing a novel fiber-optic imaging system with a scintillator to detect both 18F and fluorescent glucose probes, and (2 validating the system on ex vivo murine plaques.A novel design implements a flexible fiber-optic catheter consisting of both a radio-luminescence and a fluorescence imaging system to detect radionuclide 18F-fluorodeoxyglucose (18F-FDG and the fluorescent analog 6-(N-(7-Nitrobenz-2-oxa-1,3-diazol-4-ylamino-6-Deoxyglucose (6-NBDG, respectively. Murine macrophage-rich atherosclerotic carotid plaques were imaged ex vivo after intravenous delivery of 18F-FDG or 6-NBDG. Confirmatory optical imaging by IVIS-200 and autoradiography were also performed.Our fiber-optic imaging system successfully visualized both 18F-FDG and 6-NBDG probes in atherosclerotic plaques. For 18F-FDG, the ligated left carotid arteries (LCs exhibited 4.9-fold higher radioluminescence signal intensity compared to the non-ligated right carotid arteries (RCs (2.6 × 10(4 ± 1.4 × 10(3 vs. 5.4 × 10(3 ± 1.3 × 10(3 A.U., P = 0.008. Similarly, for 6-NBDG, the ligated LCs emitted 4.3-fold brighter fluorescent signals than the control RCs (1.6 × 10(2 ± 2.7 × 10(1 vs. 3.8 × 10(1 ± 5.9 A.U., P = 0.002. The higher uptake of both 18F-FDG and 6-NBDG in ligated LCs were confirmed with the IVIS-200 system. Autoradiography further verified the higher uptake of 18F-FDG by the LCs.This novel fiber-optic imaging system was sensitive to both radionuclide and fluorescent glucose probes taken up by murine atherosclerotic plaques. In addition, 6-NBDG is a

  14. Microarray of DNA probes on carboxylate functional beads surface

    Institute of Scientific and Technical Information of China (English)

    黄承志; 李原芳; 黄新华; 范美坤

    2000-01-01

    The microarray of DNA probes with 5’ -NH2 and 5’ -Tex/3’ -NH2 modified terminus on 10 um carboxylate functional beads surface in the presence of 1-ethyl-3-(3-dimethylaminopropyl)-carbodiimide (EDC) is characterized in the preseni paper. it was found that the microarray capacity of DNA probes on the beads surface depends on the pH of the aqueous solution, the concentra-tion of DNA probe and the total surface area of the beads. On optimal conditions, the minimum distance of 20 mer single-stranded DNA probe microarrayed on beads surface is about 14 nm, while that of 20 mer double-stranded DNA probes is about 27 nm. If the probe length increases from 20 mer to 35 mer, its microarray density decreases correspondingly. Mechanism study shows that the binding mode of DNA probes on the beads surface is nearly parallel to the beads surface.

  15. Microarray of DNA probes on carboxylate functional beads surface

    Institute of Scientific and Technical Information of China (English)

    2000-01-01

    The microarray of DNA probes with 5′-NH2 and 5′-Tex/3′-NH2 modified terminus on 10 m m carboxylate functional beads surface in the presence of 1-ethyl-3-(3-dimethylaminopropyl)- carbodiimide (EDC) is characterized in the present paper. It was found that the microarray capacity of DNA probes on the beads surface depends on the pH of the aqueous solution, the concentration of DNA probe and the total surface area of the beads. On optimal conditions, the minimum distance of 20 mer single-stranded DNA probe microarrayed on beads surface is about 14 nm, while that of 20 mer double-stranded DNA probes is about 27 nm. If the probe length increases from 20 mer to 35 mer, its microarray density decreases correspondingly. Mechanism study shows that the binding mode of DNA probes on the beads surface is nearly parallel to the beads surface.

  16. Directly labeled fluorescent DNA probes for chromosome mapping

    Energy Technology Data Exchange (ETDEWEB)

    Marrone, B.L.; Deaven, L.L.; Chen, D.J.; Park, Min S.; MacInnes, M.A.; Salzman, G.C.; Yoshida, T.M.

    1995-12-31

    A new strategy is briefly described for employing nucleic acid probes that are directly labeled with fluorochromes in fluorescence in situ hybridization techniques. These probes will permit the detection, quantitation, and high-precision spatial analysis of multiple DNA sequences along a single chromosome using video-enhanced fluorescence microscopy and digital image processing and analysis. Potential advantages of direct labeled DNA probes for fluorescence in situ hybridization far surpass currently available, indirect DNA probe labeling techniques in ease of use, versatility, and increased signal- to-noise ratio.

  17. Development and Beam-Shape Analysis of an Integrated Fiber-Optic Confocal Probe for High-Precision Central Thickness Measurement of Small-Radius Lenses

    Directory of Open Access Journals (Sweden)

    Boonsong Sutapun

    2015-04-01

    Full Text Available This work describes a new design of a fiber-optic confocal probe suitable for measuring the central thicknesses of small-radius optical lenses or similar objects. The proposed confocal probe utilizes an integrated camera that functions as a shape-encoded position-sensing device. The confocal signal for thickness measurement and beam-shape data for off-axis measurement can be simultaneously acquired using the proposed probe. Placing the probe’s focal point off-center relative to a sample’s vertex produces a non-circular image at the camera’s image plane that closely resembles an ellipse for small displacements. We were able to precisely position the confocal probe’s focal point relative to the vertex point of a ball lens with a radius of 2.5 mm, with a lateral resolution of 1.2 µm. The reflected beam shape based on partial blocking by an aperture was analyzed and verified experimentally. The proposed confocal probe offers a low-cost, high-precision technique, an alternative to a high-cost three-dimensional surface profiler, for tight quality control of small optical lenses during the manufacturing process.

  18. Detection of hydrofluoric acid by a SiO2 sol-gel coating fiber-optic probe based on reflection-based localized surface plasmon resonance.

    Science.gov (United States)

    Chen, I-Cherng; Lin, Shiu-Shiung; Lin, Tsao-Jen; Du, Je-Kang

    2011-01-01

    A novel fiber-optic probe based on reflection-based localized surface plasmon resonance (LSPR) was developed to quantify the concentration of hydrofluoric acid (HF) in aqueous solutions. The LSPR sensor was constructed with a gold nanoparticle-modified PMMA fiber, integrated with a SiO(2) sol-gel coating. This fiber-sensor was utilized to assess the relationship between HF concentration and SiO(2) sol-gel layer etching reduction. The results demonstrated the LSPR sensor was capable of detecting HF-related erosion of hydrofluoric acid solutions of concentrations ranging from 1% to 5% using Relative RI Change Rates. The development of the LSPR sensor constitutes the basis of a detector with significant sensitivity for practical use in monitoring HF solution concentrations.

  19. Empirical evaluation of oligonucleotide probe selection for DNA microarrays.

    Directory of Open Access Journals (Sweden)

    Jennifer G Mulle

    Full Text Available DNA-based microarrays are increasingly central to biomedical research. Selecting oligonucleotide sequences that will behave consistently across experiments is essential to the design, production and performance of DNA microarrays. Here our aim was to improve on probe design parameters by empirically and systematically evaluating probe performance in a multivariate context. We used experimental data from 19 array CGH hybridizations to assess the probe performance of 385,474 probes tiled in the Duchenne muscular dystrophy (DMD region of the X chromosome. Our results demonstrate that probe melting temperature, single nucleotide polymorphisms (SNPs, and homocytosine motifs all have a strong effect on probe behavior. These findings, when incorporated into future microarray probe selection algorithms, may improve microarray performance for a wide variety of applications.

  20. Empirical evaluation of oligonucleotide probe selection for DNA microarrays.

    Science.gov (United States)

    Mulle, Jennifer G; Patel, Viren C; Warren, Stephen T; Hegde, Madhuri R; Cutler, David J; Zwick, Michael E

    2010-03-29

    DNA-based microarrays are increasingly central to biomedical research. Selecting oligonucleotide sequences that will behave consistently across experiments is essential to the design, production and performance of DNA microarrays. Here our aim was to improve on probe design parameters by empirically and systematically evaluating probe performance in a multivariate context. We used experimental data from 19 array CGH hybridizations to assess the probe performance of 385,474 probes tiled in the Duchenne muscular dystrophy (DMD) region of the X chromosome. Our results demonstrate that probe melting temperature, single nucleotide polymorphisms (SNPs), and homocytosine motifs all have a strong effect on probe behavior. These findings, when incorporated into future microarray probe selection algorithms, may improve microarray performance for a wide variety of applications.

  1. Plasma polymerized epoxide functional surfaces for DNA probe immobilization.

    Science.gov (United States)

    Chu, Li-Qiang; Knoll, Wolfgang; Förch, Renate

    2008-09-15

    The development of functional surfaces for the immobilization of DNA probe is crucial for a successful design of a DNA sensor. In this report, epoxide functional thin films were achieved simply by pulsed plasma polymerization (PP) of glycidyl methacrylate (GMA) at low duty cycle. The presence of epoxide groups in the resulting ppGMA films was confirmed by Fourier transform infrared spectroscopy. The ppGMA coatings were found to be resistant to the non-specific adsorption of DNA strands, while the epoxide groups obtained could react with amine-modified DNA probes in a mild basic environment without any activation steps. A DNA sensor was made, and was successfully employed to distinguish different DNA sequences with one base pair mismatch as seen by surface plasmon enhanced fluorescence spectroscopy (SPFS). The regeneration of the present DNA sensor was also discussed. This result suggests that surface modification with ppGMA films is very promising for the fabrication of various DNA sensors.

  2. DNA nanostructure-based imaging probes and drug carriers.

    Science.gov (United States)

    Zhan, Pengfei; Jiang, Qiao; Wang, Zhen-Gang; Li, Na; Yu, Haiyin; Ding, Baoquan

    2014-09-01

    Self-assembled DNA nanostructures are well-defined nanoscale shapes, with uniform sizes, precise spatial addressability, and excellent biocompatibility. With these features, DNA nanostructures show great potential for biomedical applications; various DNA-based biomedical imaging probes or payload delivery carriers have been developed. In this review, we summarize the recent developments of DNA-based nanostructures as tools for diagnosis and cancer therapy. The biological effects that are brought about by DNA nanostructures are highlighted by in vitro and in vivo imaging, antitumor drug delivery, and immunostimulatory therapy. The challenges and perspectives of DNA nanostructures in the field of nanomedicine are discussed.

  3. The detection of HBV DNA with gold nanoparticle gene probes

    Institute of Scientific and Technical Information of China (English)

    Dong Xi; Xiaoping Luo; Qin Ning; Qianghua Lu; Kailun Yao; Zuli Liu

    2007-01-01

    Objective:Gold nanoparticle Hepatitis B virus (HBV) DNA probes were prepared, and their application for HBV DNA measurement was studied. Methods:Alkanethiol modified oligonucleotide was bound with self-made Au nanoparticles to form nanoparticle HBV DNA gene probes, through covalent binding of Au-S. By using a fluorescence-based method, the number of thiol-derivatized, single-stranded oligonucleotides and their hybridization efficiency with complementary oligonucleotides in solution was determined. With the aid of Au nanoparticle-supported mercapto-modified oligonucleotides serving as detection probes, and oligonucleotides immobilized on a nylon membrane surface acting as capturing probes,HBV DNA was detected visually by sandwich hybridization based on highly sensitive aggregation and silver staining. The modified nanoparticle HBV DNA gene probes were also used to detect the HBV DNA extracted from serum in patients with hepatitis B. Results:Compared with bare Au nanoparticles, oligonucleotide modified nanoparticles had a higher stability in NaCl solution or under high temperature environment and the absorbance peak of modified Au nanoparticles shifted from 520nm to 524nm. For Au nanoparticles, the maximal oligonucleotide surface coverage of hexaethiol 30-mer oligonucleotide was (132 ± 10) oligonucleotides per nanoparticle, and the percentage of hybridization strands on nanoparticles was (22 ± 3% ). Based on a two-probe sandwich hybridization/nanoparticle amplification/silver staining enhancement method, Au nanoparticle gene probes could detect as low as 10-11 mol/L composite HBV DNA molecules on a nylon membrane and the PCR products of HBV DNA visually. As made evident by transmission electron microscopy, the nanoparticles assembled into large network aggregates when nanoparticle HBV DNA gene probes were applied to detect HBV DNA molecules in liquid. Conclusion:Our results showed that successfully prepared Au nanoparticle HBV DNA gene probes could be used to

  4. Fluorescent cyanine probe for DNA detection and cellular imaging

    Science.gov (United States)

    Zheng, Yong-Chao; Zheng, Mei-Ling; Zhao, Zhen-Sheng; Duan, Xuan-Ming

    2014-03-01

    In our study, two carbazole-based cyanines, 3,6-bis[2-(1-methylpyridinium)vinyl]-9-methyl carbazole diiodide (A) and 6,6'-bis[2-(1-methylpyridinium)vinyl]-bis(9-methyl-carbazol-3yl)methane diiodide (B) were synthesized and employed as light-up probes for DNA and cell imaging. Both of the cyanine probes possess a symmetric structure and bis-cationic center. The obvious induced circular dichroism signals in circular dichroism spectra reveal that the molecules can specifically interact with DNA. Strong fluorescence enhancement is observed when these two cyanines are bound to DNA. These cyanine probes show high binding affinity to oligonucleotides but different binding preferences to various secondary structures. Confocal microscopy images of fixed cell stained by the probes exhibit strong brightness and high contrast in nucleus with a very low cytoplasmic background.

  5. Utilizing Gold Nanoparticle Probes to Visually Detect DNA Methylation

    Science.gov (United States)

    Chen, Kui; Zhang, Mingyi; Chang, Ya-Nan; Xia, Lin; Gu, Weihong; Qin, Yanxia; Li, Juan; Cui, Suxia; Xing, Gengmei

    2016-06-01

    The surface plasmon resonance (SPR) effect endows gold nanoparticles (GNPs) with the ability to visualize biomolecules. In the present study, we designed and constructed a GNP probe to allow the semi-quantitative analysis of methylated tumor suppressor genes in cultured cells. To construct the probe, the GNP surfaces were coated with single-stranded DNA (ssDNA) by forming Au-S bonds. The ssDNA contains a thiolated 5'-end, a regulatory domain of 12 adenine nucleotides, and a functional domain with absolute pairing with methylated p16 sequence (Met- p16). The probe, paired with Met- p16, clearly changed the color of aggregating GNPs probe in 5 mol/L NaCl solution. Utilizing the probe, p16 gene methylation in HCT116 cells was semi-quantified. Further, the methylation of E-cadherin, p15, and p16 gene in Caco2, HepG2, and HCT116 cell lines were detected by the corresponding probes, constructed with three domains. This simple and cost-effective method was useful for the diagnosis of DNA methylation-related diseases.

  6. Probing the elastic limit of DNA bending

    CERN Document Server

    Le, Tung T

    2014-01-01

    Many structures inside the cell such as nucleosomes and protein-mediated DNA loops contain sharply bent double-stranded (ds) DNA. Therefore, the energetics of strong dsDNA bending constitutes an essential part of cellular thermodynamics. Although the thermomechanical behavior of long dsDNA is well described by the worm-like chain (WLC) model, the length limit of such elastic behavior remains controversial. To investigate the energetics of strong dsDNA bending, we measured the opening rate of small dsDNA loops with contour lengths of 40-200 bp using Fluorescence Resonance Energy Transfer (FRET). From the measured relationship of loop stability to loop size, we observed a transition between two separate bending regimes at a critical loop size below 100 bp. Above this loop size, the loop lifetime decreased with decreasing loop size in a manner consistent with an elastic bending stress. Below the critical loop size, however, the loop lifetime became less sensitive to loop size, indicative of softening of the doub...

  7. Effects of nicotinamide and carbogen on oxygenation in human tumor xenografts measured with luminescense based fiber-optic probes.

    Science.gov (United States)

    Bussink, J; Kaanders, J H; Strik, A M; van der Kogel, A J

    2000-10-01

    In head and neck cancer, addition of both carbogen breathing and nicotinamide to accelerated fractionated radiotherapy showed increased loco-regional control rates. An assay based on the measurement of changes in tumor pO(2) in response to oxygenation modification could be helpful for selecting patients for these new treatment approaches. The fiber-optic oxygen-sensing device, OxyLite, was used to measure changes in pO(2), at a single position in tumors, after treatment with nicotinamide and carbogen in three human xenograft tumor lines with different vascular architecture and hypoxic patterns. Pimonidazole was used as a marker of hypoxia and was analyzed with a digital image processing system. At the position of pO(2) measurement, half of the tumors showed a local increase in pO(2) after nicotinamide administration. Steep increases in pO(2) were measured in most tumors during carbogen breathing although the increase was less pronounced in tumor areas with a low pre-treatment pO(2). A trend towards a faster local response to carbogen breathing for nicotinamide pre-treated tumors was found in all three lines. There were significant differences in hypoxic fractions, based on pimonidazole binding, between the three tumor lines. There was no correlation between hypoxic marker binding and the response to carbogen breathing. Temporal changes in local pO(2) can be measured with the OxyLite. This system was used to quantitate the effects of oxygen modifying treatments. Rapid increases in pO(2) during carbogen breathing were observed in most tumor areas. The locally measured response to nicotinamide was smaller and more variable. Bio-reductive hypoxic cell marker binding in combination with OxyLite pO(2) determination gives spatial information about the distribution patterns of tumor hypoxia at the microscopic level together with the possibility to continuously measure changes in pO(2) in specific tumor areas.

  8. Probe Selection for DNA Microarrays using OligoWiz

    DEFF Research Database (Denmark)

    Wernersson, Rasmus; Juncker, Agnieszka; Nielsen, Henrik Bjørn

    2007-01-01

    Nucleotide abundance measurements using DNA microarray technology are possible only if appropriate probes complementary to the target nucleotides can be identified. Here we present a protocol for selecting DNA probes for microarrays using the OligoWiz application. OligoWiz is a client......-server application that offers a detailed graphical interface and real-time user interaction on the client side, and massive computer power and a large collection of species databases (400, summer 2007) on the server side. Probes are selected according to five weighted scores: cross-hybridization, deltaT(m), folding...... computer skills and can be executed from any Internet-connected computer. The probe selection procedure for a standard microarray design targeting all yeast transcripts can be completed in 1 h....

  9. Colorimetric DNA detection of transgenic plants using gold nanoparticles functionalized with L-shaped DNA probes

    Science.gov (United States)

    Nourisaeid, Elham; Mousavi, Amir; Arpanaei, Ayyoob

    2016-01-01

    In this study, a DNA colorimetric detection system based on gold nanoparticles functionalized with L-shaped DNA probes was prepared and evaluated. We investigated the hybridization efficiency of the L-shaped probes and studied the effect of nanoparticle size and the L-shaped DNA probe length on the performance of the as-prepared system. Probes were attached to the surface of gold nanoparticles using an adenine sequence. An optimal sequence of 35S rRNA gene promoter from the cauliflower mosaic virus, which is frequently used in the development of transgenic plants, and the two complementary ends of this gene were employed as model target strands and probe molecules, respectively. The spectrophotometric properties of the as-prepared systems indicated that the large NPs show better changes in the absorption spectrum and consequently present a better performance. The results of this study revealed that the probe/Au-NPs prepared using a vertical spacer containing 5 thymine oligonucleotides exhibited a stronger spectrophotometric response in comparison to that of larger probes. These results in general indicate the suitable performance of the L-shaped DNA probe-functionalized Au-NPs, and in particular emphasize the important role of the gold nanoparticle size and length of the DNA probes in enhancing the performance of such a system.

  10. Multilayers Assembly of DNA Probe for Biosensor

    Institute of Scientific and Technical Information of China (English)

    谢文章; 路英杰; 隋森芳

    2002-01-01

    Surface plasmon resonance (SPR) was a sensitive method to study molecular interactions. Based on the specific binding, this paper presented the molecular assembly of protein-nucleic acid multilayers on the surface of a gold film. The first layer was a biotin-lipid (B-DMPE/DMPE) containing a monolayer prepared using the Langmuir-Blodgett (LB) technique. The second and third layers were avidin and DNA labeled biotin, respectively. The fourth layer was anti-DNA antibody extracted from the serum of patients with systemic lupus erythematosus (SLE). These interactions provide stability in the multilayer films of the complexes. The multilayer formation process was detected by SPR spectroscopy. The results show that the chip-based sensor system can be used for functional characterization of protein-protein and protein-DNA interactions.

  11. Probing the microscopic flexibility of DNA from melting temperatures

    Science.gov (United States)

    Weber, Gerald; Essex, Jonathan W.; Neylon, Cameron

    2009-10-01

    The microscopic flexibility of DNA is a key ingredient for understanding its interaction with proteins and drugs but is still poorly understood and technically challenging to measure. Several experimental methods probe very long DNA samples, but these miss local flexibility details. Others mechanically disturb or modify short molecules and therefore do not obtain flexibility properties of unperturbed and pristine DNA. Here, we show that it is possible to extract very detailed flexibility information about unmodified DNA from melting temperatures with statistical physics models. We were able to retrieve, from published melting temperatures, several established flexibility properties such as the presence of highly flexible TATA regions of genomic DNA and support recent findings that DNA is very flexible at short length scales. New information about the nanoscale Na+ concentration dependence of DNA flexibility was determined and we show the key role of ApT and TpA steps when it comes to ion-dependent flexibility and melting temperatures.

  12. Evolving DNA motifs to predict GeneChip probe performance

    Directory of Open Access Journals (Sweden)

    Harrison AP

    2009-03-01

    Full Text Available Abstract Background Affymetrix High Density Oligonuclotide Arrays (HDONA simultaneously measure expression of thousands of genes using millions of probes. We use correlations between measurements for the same gene across 6685 human tissue samples from NCBI's GEO database to indicated the quality of individual HG-U133A probes. Low correlation indicates a poor probe. Results Regular expressions can be automatically created from a Backus-Naur form (BNF context-free grammar using strongly typed genetic programming. Conclusion The automatically produced motif is better at predicting poor DNA sequences than an existing human generated RE, suggesting runs of Cytosine and Guanine and mixtures should all be avoided.

  13. A new method of preparing fiber-optic DNA biosensor and its array for gene detection

    Institute of Scientific and Technical Information of China (English)

    JIANG; Guangfen; (

    2001-01-01

    , 1146(1): 136.[12] Howlett, N. G., Avery, S. V., Relationship between cadmium sensitivity and degree of plasma membrane fatty acid unsatu-ration in Saccharomyces cerevisiae, Appl. Microbiol. Biotechnol., 1997, 48(4): 539.[13] Petriz, J., Oconnor, J. E., Carmona, M. et al., Is Rhodamine-123 an appropriate fluorescent probe to assess P-glycoprotein mediated multidrug resistance in vinblastine-resistant CHO cells? Analytical Cellular Pathology, 1997, 14(3): 129.[14] Leonce, S., Burbridge, M., Flow cytometry: a useful technique in the study of multidrug resistance, J. Bio. Cell, 1993, 78(1-2): 63.[15] Le Moyec, L., Tatoud, R., Degorges, A. et al., Proton nuclear magnetic resonance spectroscopy reveals cellular lipids in-volved in resistance to Adriamycin and Taxol by the K562 Leukemia cell line, Cancer Res., 1996, 56: 3461.[16] Callaghan, R., Stafford, A., Epand, R. M., Increased accumulation of drugs in a multidrug resistant cell line by alteration of membrane biophysical properties, Biochim. Biophys. Acta, 1993, 1175(3): 277.[17] Sinicrope, F. A., Dudeia, P. K., Bissommette, B. M. et al., Modulation of P-glycoprotein-mediated drug transport by al-terations in lipid fluidity of rat liver canlicular membrane vesicles, J. Biol. Chem., 1992, 267(35): 24995.[18] Romsicki, Y., Sharom, F. J., The membrane lipid environment modulates drug interactions with the P-glycoprotein multi-drug transporter, Biochemistry, 1999, 38(21): 6887.[19] Garel, O., Lecureur, V., Guillouzo, A., The P-glycoprotein multidrug transporter, Gen. Pharmacol., 1996, 27(8): 1283.[20] Aran, J. M., Pastan, I., Gottesman, M. M., Therapeutic strategies involving the multidrug resistance phenotype: the MDR1 gene as target, chemoprotectant, and selectable marker in gene therapy, Adv. Pharmacol., 1999, 46: 1.[21] Zaman, G. J., Flens, M. J., Vanleusden, M. R. et al., The human multidrug resistance-associated protein (MRP) is a plasma membrane drug efflux pump, Proc. Natl. Acad

  14. Isolation and characterization of DNA probes for human chromosome 21.

    Science.gov (United States)

    Watkins, P C

    1990-01-01

    A coordinated effort to map and sequence the human genome has recently become a national priority. Chromosome 21, the smallest human chromosome accounting for less than 2% of the human genome, is an attractive model system for developing and evaluating genome mapping technology. Several strategies are currently being explored including the development of chromosome 21 libraries from somatic cell hybrids as reported here, the cloning of chromosome 21 in yeast artificial chromosomes (McCormick et al., 1989b), and the construction of chromosome 21 libraries using chromosome flow-sorting techniques (Fuscoe et al., 1989). This report describes the approaches used to identify DNA probes that are useful for mapping chromosome 21. Probes were successfully isolated from both phage and cosmid libraries made from two somatic cell hybrids that contain human chromosome 21 as the only human chromosome. The 15 cosmid clones from the WA17 library, reduced to cloned DNA sequences of an average size of 3 kb, total 525 kb of DNA which is approximately 1% of chromosome 21. From these clones, a set of polymorphic DNA markers that span the length of the long arm of chromosome 21 has been generated. All of the probes thus far analyzed from the WA17 libraries have been mapped to chromosome 21 both by physical and genetic mapping methods. It is therefore likely that the WA17 hybrid cell line contains human chromosome 21 as the only human component, in agreement with cytogenetic observation. The 153E7b cosmid libraries will provide an alternative source of cloned chromosome 21 DNA. Library screening techniques can be employed to obtain cloned DNA sequences from the same genetic loci of the two different chromosome 21s. Comparative analysis will allow direct estimation of DNA sequence variation for different regions of chromosome 21. Mapped DNA probes make possible the molecular analysis of chromosome 21 at a level of resolution not achievable by classical cytogenetic techniques (Graw et al

  15. Ultrasonic manipulation of yeast cells in suspension for absorption spectroscopy with an immersible mid-infrared fiberoptic probe.

    Science.gov (United States)

    Koch, Cosima; Brandstetter, Markus; Lendl, Bernhard; Radel, Stefan

    2013-06-01

    Recent advances in combining ultrasonic particle manipulation with attenuated total reflection infrared spectroscopy of yeast suspensions are presented. Infrared spectroscopy provides highly specific molecular information about the sample. It has not been applicable to in-line monitoring of cells during fermentation, however, because positioning cells in the micron-thin measurement region of the attenuated total reflection probe was not possible. Ultrasonic radiation forces exerted on suspended particles by an ultrasonic standing wave can result in the buildup of agglomerates in the nodal planes, hence enabling the manipulation of suspended cells on the microscopic scale. When a chamber setup and a prototype in-line applicable probe were used, successful control over the position of the yeast cells relative to the attenuated total reflection sensor surface could be proven. Both rate of increase and maximum mid-infrared absorption of yeast-specific bands during application of a pushing frequency (chamber setup: 1.863 MHz, in-line probe: 1.990 MHz) were found to correlate with yeast cell concentration.

  16. Probe Microscopic Studies of DNA Molecules on Carbon Nanotubes

    Directory of Open Access Journals (Sweden)

    Kazuo Umemura

    2016-10-01

    Full Text Available Hybrids of DNA and carbon nanotubes (CNTs are promising nanobioconjugates for nanobiosensors, carriers for drug delivery, and other biological applications. In this review, nanoscopic characterization of DNA-CNT hybrids, in particular, characterization by scanning probe microscopy (SPM, is summarized. In many studies, topographical imaging by atomic force microscopy has been performed. However, some researchers have demonstrated advanced SPM operations in order to maximize its unique and valuable functions. Such sophisticated approaches are attractive and will have a significant impact on future studies of DNA-CNT hybrids.

  17. Probing Nucleosome Stability with a DNA Origami Nanocaliper.

    Science.gov (United States)

    Le, Jenny V; Luo, Yi; Darcy, Michael A; Lucas, Christopher R; Goodwin, Michelle F; Poirier, Michael G; Castro, Carlos E

    2016-07-26

    The organization of eukaryotic DNA into nucleosomes and chromatin undergoes dynamic structural changes to regulate genome processing, including transcription and DNA repair. Critical chromatin rearrangements occur over a wide range of distances, including the mesoscopic length scale of tens of nanometers. However, there is a lack of methodologies that probe changes over this mesoscopic length scale within chromatin. We have designed, constructed, and implemented a DNA-based nanocaliper that probes this mesoscopic length scale. We developed an approach of integrating nucleosomes into our nanocaliper at two attachment points with over 50% efficiency. Here, we focused on attaching the two DNA ends of the nucleosome to the ends of the two nanocaliper arms, so the hinge angle is a readout of the nucleosome end-to-end distance. We demonstrate that nucleosomes integrated with 6, 26, and 51 bp linker DNA are partially unwrapped by the nanocaliper by an amount consistent with previously observed structural transitions. In contrast, the nucleosomes integrated with the longer 75 bp linker DNA remain fully wrapped. We found that the nanocaliper angle is a sensitive measure of nucleosome disassembly and can read out transcription factor (TF) binding to its target site within the nucleosome. Interestingly, the nanocaliper not only detects TF binding but also significantly increases the probability of TF occupancy at its site by partially unwrapping the nucleosome. These studies demonstrate the feasibility of using DNA nanotechnology to both detect and manipulate nucleosome structure, which provides a foundation of future mesoscale studies of nucleosome and chromatin structural dynamics.

  18. Intra-Tissue Pressure Measurement in Ex Vivo Liver Undergoing Laser Ablation with Fiber-Optic Fabry-Perot Probe.

    Science.gov (United States)

    Tosi, Daniele; Saccomandi, Paola; Schena, Emiliano; Duraibabu, Dinesh Babu; Poeggel, Sven; Leen, Gabriel; Lewis, Elfed

    2016-04-15

    We report the first-ever intra-tissue pressure measurement performed during 1064 nm laser ablation (LA) of an ex vivo porcine liver. Pressure detection has been performed with a biocompatible, all-glass, temperature-insensitive Extrinsic Fabry-Perot Interferometry (EFPI) miniature probe; the proposed methodology mimics in-vivo treatment. Four experiments have been performed, positioning the probe at different positions from the laser applicator tip (from 0.5 mm to 5 mm). Pressure levels increase during ablation time, and decrease with distance from applicator tip: the recorded peak parenchymal pressure levels range from 1.9 kPa to 71.6 kPa. Different pressure evolutions have been recorded, as pressure rises earlier in proximity of the tip. The present study is the first investigation of parenchymal pressure detection in liver undergoing LA: the successful detection of intra-tissue pressure may be a key asset for improving LA, as pressure levels have been correlated to scattered recurrences of tumors by different studies.

  19. Intra-Tissue Pressure Measurement in Ex Vivo Liver Undergoing Laser Ablation with Fiber-Optic Fabry-Perot Probe

    Directory of Open Access Journals (Sweden)

    Daniele Tosi

    2016-04-01

    Full Text Available We report the first-ever intra-tissue pressure measurement performed during 1064 nm laser ablation (LA of an ex vivo porcine liver. Pressure detection has been performed with a biocompatible, all-glass, temperature-insensitive Extrinsic Fabry-Perot Interferometry (EFPI miniature probe; the proposed methodology mimics in-vivo treatment. Four experiments have been performed, positioning the probe at different positions from the laser applicator tip (from 0.5 mm to 5 mm. Pressure levels increase during ablation time, and decrease with distance from applicator tip: the recorded peak parenchymal pressure levels range from 1.9 kPa to 71.6 kPa. Different pressure evolutions have been recorded, as pressure rises earlier in proximity of the tip. The present study is the first investigation of parenchymal pressure detection in liver undergoing LA: the successful detection of intra-tissue pressure may be a key asset for improving LA, as pressure levels have been correlated to scattered recurrences of tumors by different studies.

  20. A simple and rapid method for the preparation of homologous DNA oligonucleotide hybridization probes from heterologous gene sequences and probes.

    Science.gov (United States)

    Maxwell, E S; Sarge, K D

    1988-11-30

    We describe a simple and rapid method for the preparation of homologous DNA oligonucleotide probes for hybridization analysis and/or cDNA/genomic library screening. With this method, a synthetic DNA oligonucleotide derived from a known heterologous DNA/RNA/protein sequence is annealed to an RNA preparation containing the gene transcript of interest. Any unpaired 3'-terminal oligonucleotides of the heterologous DNA primer are then removed using the 3' exonuclease activity of the DNA Polymerase I Klenow fragment before primer extension/dideoxynucleotide sequencing of the annealed RNA species with AMV reverse transcriptase. From the determined RNA sequence, a completely homologous DNA oligonucleotide probe is then prepared. This approach has been used to prepare a homologous DNA oligonucleotide probe for the successful library screening of the yeast hybRNA gene starting with a heterologous mouse hybRNA DNA oligonucleotide probe.

  1. DNA Probe Pooling for Rapid Delineation of Chromosomal Breakpoints

    Energy Technology Data Exchange (ETDEWEB)

    Lu, Chun-Mei; Kwan, Johnson; Baumgartner, Adolf; Weier, Jingly F.; Wang, Mei; Escudero, Tomas; Munne' , Santiago; Zitzelsberger, Horst F.; Weier, Heinz-Ulrich

    2009-01-30

    Structural chromosome aberrations are hallmarks of many human genetic diseases. The precise mapping of translocation breakpoints in tumors is important for identification of genes with altered levels of expression, prediction of tumor progression, therapy response, or length of disease-free survival as well as the preparation of probes for detection of tumor cells in peripheral blood. Similarly, in vitro fertilization (IVF) and preimplantation genetic diagnosis (PGD) for carriers of balanced, reciprocal translocations benefit from accurate breakpoint maps in the preparation of patient-specific DNA probes followed by a selection of normal or balanced oocytes or embryos. We expedited the process of breakpoint mapping and preparation of case-specific probes by utilizing physically mapped bacterial artificial chromosome (BAC) clones. Historically, breakpoint mapping is based on the definition of the smallest interval between proximal and distal probes. Thus, many of the DNA probes prepared for multi-clone and multi-color mapping experiments do not generate additional information. Our pooling protocol described here with examples from thyroid cancer research and PGD accelerates the delineation of translocation breakpoints without sacrificing resolution. The turnaround time from clone selection to mapping results using tumor or IVF patient samples can be as short as three to four days.

  2. Validation of DNA probes for molecular cytogenetics by mapping onto immobilized circular DNA

    Energy Technology Data Exchange (ETDEWEB)

    Greulich-Bode, Karin M.; Wang, Mei; Rhein, Andreas P.; Weier, Jingly F.; Weier, Heinz-Ulli G.

    2008-12-04

    Fluorescence in situ hybridization (FISH) is a sensitive and rapid procedure to detect gene rearrangements in tumor cells using non-isotopically labeled DNA probes. Large insert recombinant DNA clones such as bacterial artificial chromosome (BAC) or P1/PAC clones have established themselves in recent years as preferred starting material for probe preparations due to their low rates of chimerism and ease of use. However, when developing probes for the quantitative analysis of rearrangements involving genomic intervals of less than 100kb, careful probe selection and characterization are of paramount importance. We describe a sensitive approach to quality control probe clones suspected of carrying deletions or for measuring clone overlap with near kilobase resolution. The method takes advantage of the fact that P1/PAC/BAC's can be isolated as circular DNA molecules, stretched out on glass slides and fine-mapped by multicolor hybridization with smaller probe molecules. Two examples demonstrate the application of this technique: mapping of a gene-specific {approx}6kb plasmid onto an unusually small, {approx}55kb circular P1 molecule and the determination of the extent of overlap between P1 molecules homologous to the human NF-{kappa}B2 locus. The relatively simple method presented here does not require specialized equipment and may thus find widespread applications in DNA probe preparation and characterization, the assembly of physical maps for model organisms or in studies on gene rearrangements.

  3. Validation of DNA probes for molecular cytogenetics by mapping onto immobilized circular DNA

    Energy Technology Data Exchange (ETDEWEB)

    Greulich-Bode, Karin; Wang, Mei; Rhein, Andreas; Weier, Jingly; Weier, Heinz-Ulli

    2008-12-16

    Fluorescence in situ hybridization (FISH) is a sensitive and rapid procedure to detect gene rearrangements in tumor cells using non-isotopically labeled DNA probes. Large insert recombinant DNA clones such as bacterial artificial chromosome (BAC) or P1/PAC clones have established themselves in recent years as preferred starting material for probe preparations due to their low rates of chimerism and ease of use. However, when developing probes for the quantitative analysis of rearrangements involving genomic intervals of less than 100kb, careful probe selection and characterization are of paramount importance. We describe a sensitive approach to quality control probe clones suspected of carrying deletions or for measuring clone overlap with near kilobase resolution. The method takes advantage of the fact that P1/PAC/BAC's can be isolated as circular DNA molecules, stretched out on glass slides and fine-mapped by multicolor hybridization with smaller probe molecules. Two examples demonstrate the application of this technique: mapping of a gene-specific {approx}6kb plasmid onto an unusually small, {approx}55kb circular P1 molecule and the determination of the extent of overlap between P1 molecules homologous to the human NF-?B2 locus. The relatively simple method presented here does not require specialized equipment and may thus find widespread applications in DNA probe preparation and characterization, the assembly of physical maps for model organisms or in studies on gene rearrangements.

  4. Validation of DNA probes for molecular cytogenetics by mapping onto immobilized circular DNA

    Directory of Open Access Journals (Sweden)

    Rhein Andreas P

    2008-12-01

    Full Text Available Abstract Background Fluorescence in situ hybridization (FISH is a sensitive and rapid procedure to detect gene rearrangements in tumor cells using non-isotopically labeled DNA probes. Large insert recombinant DNA clones such as bacterial artificial chromosome (BAC or P1/PAC clones have established themselves in recent years as preferred starting material for probe preparations due to their low rates of chimerism and ease of use. However, when developing probes for the quantitative analysis of rearrangements involving genomic intervals of less than 100 kb, careful probe selection and characterization are of paramount importance. Results We describe a sensitive approach to quality control probe clones suspected of carrying deletions or for measuring clone overlap with near kilobase resolution. The method takes advantage of the fact that P1/PAC/BAC's can be isolated as circular DNA molecules, stretched out on glass slides and fine-mapped by multicolor hybridization with smaller probe molecules. Two examples demonstrate the application of this technique: mapping of a gene-specific ~6 kb plasmid onto an unusually small, ~55 kb circular P1 molecule and the determination of the extent of overlap between P1 molecules homologous to the human NF-κB2 locus. Conclusion The relatively simple method presented here does not require specialized equipment and may thus find widespread applications in DNA probe preparation and characterization, the assembly of physical maps for model organisms or in studies on gene rearrangements.

  5. Mobile fiber-optic laser Doppler anemometer.

    Science.gov (United States)

    Stieglmeier, M; Tropea, C

    1992-07-20

    A laser Doppler anemometer (LDA) has been developed that combines the compactness and low power consumption of laser diodes and avalanche photodiodes with the flexibility and possibility of miniaturization by using fiber-optic probes. The system has been named DFLDA for laser diode fiber LDA and is especially suited for mobile applications, for example, in trains, airplanes, or automobiles. Optimization considerations of fiber-optic probes are put forward and several probe examples are described in detail. Measurement results from three typical applications are given to illustrate the use of the DFLDA. Finally, a number of future configurations of the DFLDA concept are discussed.

  6. Chemical Biology Probes from Advanced DNA-encoded Libraries.

    Science.gov (United States)

    Salamon, Hazem; Klika Škopić, Mateja; Jung, Kathrin; Bugain, Olivia; Brunschweiger, Andreas

    2016-02-19

    The identification of bioactive compounds is a crucial step toward development of probes for chemical biology studies. Screening of DNA-encoded small molecule libraries (DELs) has emerged as a validated technology to interrogate vast chemical space. DELs consist of chimeric molecules composed of a low-molecular weight compound that is conjugated to a DNA identifier tag. They are screened as pooled libraries using selection to identify "hits." Screening of DELs has identified numerous bioactive compounds. Some of these molecules were instrumental in gaining a deeper understanding of biological systems. One of the main challenges in the field is the development of synthesis methodology for DELs.

  7. Photoenzyme probes of photodamage to cells and cellular DNA

    Energy Technology Data Exchange (ETDEWEB)

    Sutherland, B. M.

    1979-01-01

    Development of photoenzyme probes for detection of ultraviolet damage to cells and DNA is reviewed with special emphasis on a process using polyethylene glycol to induce cell fusion. Polyethylene glycol is easy to obtain and handle, is gentle to the cells and does not induce latent or productive virus infection; therefore, it may be a general method for insertion of exogenous enzymes into mammalian cells. (PCS)

  8. Label-Free Potentiometry for Detecting DNA Hybridization Using Peptide Nucleic Acid and DNA Probes

    Directory of Open Access Journals (Sweden)

    Yuji Miyahara

    2013-02-01

    Full Text Available Peptide nucleic acid (PNA has outstanding affinity over DNA for complementary nucleic acid sequences by forming a PNA-DNA heterodimer upon hybridization via Watson-Crick base-pairing. To verify whether PNA probes on an electrode surface enhance sensitivity for potentiometric DNA detection or not, we conducted a comparative study on the hybridization of PNA and DNA probes on the surface of a 10-channel gold electrodes microarray. Changes in the charge density as a result of hybridization at the solution/electrode interface on the self-assembled monolayer (SAM-formed microelectrodes were directly transformed into potentiometric signals using a high input impedance electrometer. The charge readout allows label-free, reagent-less, and multi-parallel detection of target oligonucleotides without any optical assistance. The differences in the probe lengths between 15- to 22-mer dramatically influenced on the sensitivity of the PNA and DNA sensors. Molecular type of the capturing probe did not affect the degree of potential shift. Theoretical model for charged rod-like duplex using the Gouy-Chapman equation indicates the dominant effect of electrostatic attractive forces between anionic DNA and underlying electrode at the electrolyte/electrode interface in the potentiometry.

  9. Label-free potentiometry for detecting DNA hybridization using peptide nucleic acid and DNA probes.

    Science.gov (United States)

    Goda, Tatsuro; Singi, Ankit Balram; Maeda, Yasuhiro; Matsumoto, Akira; Torimura, Masaki; Aoki, Hiroshi; Miyahara, Yuji

    2013-02-07

    Peptide nucleic acid (PNA) has outstanding affinity over DNA for complementary nucleic acid sequences by forming a PNA-DNA heterodimer upon hybridization via Watson-Crick base-pairing. To verify whether PNA probes on an electrode surface enhance sensitivity for potentiometric DNA detection or not, we conducted a comparative study on the hybridization of PNA and DNA probes on the surface of a 10-channel gold electrodes microarray. Changes in the charge density as a result of hybridization at the solution/electrode interface on the self-assembled monolayer (SAM)-formed microelectrodes were directly transformed into potentiometric signals using a high input impedance electrometer. The charge readout allows label-free, reagent-less, and multi-parallel detection of target oligonucleotides without any optical assistance. The differences in the probe lengths between 15- to 22-mer dramatically influenced on the sensitivity of the PNA and DNA sensors. Molecular type of the capturing probe did not affect the degree of potential shift. Theoretical model for charged rod-like duplex using the Gouy-Chapman equation indicates the dominant effect of electrostatic attractive forces between anionic DNA and underlying electrode at the electrolyte/electrode interface in the potentiometry.

  10. RNA probes, transcribed from synthetic DNA, for in situ hybridization

    Energy Technology Data Exchange (ETDEWEB)

    Brysch, W.; Hagendorff, G.; Schlingensiepen, K.H.

    1988-03-25

    Single stranded cRNA probes are ideal for in-situ-hybridization. Synthetic oligodesoxy-ribonucleotides on the other hand allow one to chose nucleotide sequences independently of restriction sites and availability of cloned templates. To combine the advantages of these two methods, the authors used an oligonucleotide, containing a T7-RNA-polymerase promotor sequence and a starting sequence of 6 bases as a template for an in-vitro-transcription reaction with T7-RNA-polymerase. A second oligonucleotide, complementary to basepairs 1-101 was also synthesized and the two strands heated to 95/sup 0/ for 3 min, then kept at 65/sup 0/C for one hour in 80 mM Tris, 12mM MgCl, 4 mM Spermidine, 0,04% Triton and finally cooled on ice. The resulting double stranded DNA was used as a template to transcribe /sup 35/S-labelled cRNA, using DNA, T7-Polymerase, /sup 35/S-UTP, ATP, GTP and CTP and RNasin (Promega). No difference could be observed comparing the resulting hybridization pattern with that obtained by using a plasmid derived cRNA probe of rat brain sodium channel II. Moreover the hybridization signal was clearly distinct from the background labelling resulting from hybridization with a sense control probe of the same specific activity.

  11. Isolation of human minisatellite loci detected by synthetic tandem repeat probes: direct comparison with cloned DNA fingerprinting probes.

    Science.gov (United States)

    Armour, J A; Vergnaud, G; Crosier, M; Jeffreys, A J

    1992-08-01

    As a direct comparison with cloned 'DNA fingerprinting' probes, we present the results of screening an ordered array Charomid library for hypervariable human loci using synthetic tandem repeat (STR) probes. By recording the coordinates of positive hybridization signals, the subset of clones within the library detected by each STR probe can be defined, and directly compared with the set of clones detected by naturally occurring (cloned) DNA fingerprinting probes. The STR probes vary in the efficiency of detection of polymorphic minisatellite loci; among the more efficient probes, there is a strong overlap with the sets of clones detected by the DNA fingerprinting probes. Four new polymorphic loci were detected by one or more of the STR probes but not by any of the naturally occurring repeats. Sequence comparisons with the probe(s) used to detect the locus suggest that a relatively poor match, for example 10 out of 14 bases in a limited region of each repeat, is sufficient for the positive detection of tandem repeats in a clone in this type of library screening by hybridization. These results not only provide a detailed evaluation of the usefulness of STR probes in the isolation of highly variable loci, but also suggest strategies for the use of these multi-locus probes in screening libraries for clones from hypervariable loci.

  12. COMPARISON OF DIFFERENT ENZYMES AND PROBES AND THEIR COMBINATIONS IN DNA FINGERPRINTING

    Institute of Scientific and Technical Information of China (English)

    2001-01-01

    In the present study, eight combinations of restriction enzymes and oligonucleotide probes were tested for detecting VNTR polymorphism. More than a hundred loci were detected by all enzyme-probe combi nations. The influences of breed, enzyme and probe as well as their interactions were analysed, and the mean value of DNA fingerprint data was calculated for the enzymes and probes. The results will provide some valu- able information for studying the genetic relationship of individuals or populations using DNA fingerprinting.

  13. Probing the Structure of DNA Aptamers with a Classic Heterocycle.

    Directory of Open Access Journals (Sweden)

    G. Reid Bishop

    2004-02-01

    Full Text Available DNA aptamers are synthetic, single-stranded DNA oligonucleotides selectedby SELEX methods for their binding with specific ligands. Here we present ethidiumbinding results for three related DNA aptamers (PDB code: 1OLD, 1DB6, and 2ARGthat bind L-argininamide (L-Arm. The ligand bound form of each aptamer's structurehas been reported and each are found to be composed primarily of two domainsconsisting of a stem helical region and a loop domain that forms a binding pocket for thecognate ligand. Previous thermodynamic experiments demonstrated that the DNAaptamer 1OLD undergoes a large conformational ordering upon binding to L-Arm. Herewe extend those linkage binding studies by examining the binding of the heterocyclicintercalator ethidium to each of the three aptamers by fluorescence and absorptionspectrophotometric titrations. Our results reveal that ethidium binds to each aptamer with∆Go's in the range of -8.7 to -9.4 kcal/mol. The stoichiometry of binding is 2:1 for eachaptamer and is quantitatively diminished in the presence of L-Arm as is the overallfluorescence intensity of ethidium. Together, these results demonstrate that a portion ofthe bound ethidium is excluded from the aptamer in the presence of a saturating amountof L-Arm. These results demonstrate the utility of ethidium and related compounds forthe probing of non-conventional DNA structures and reveal an interesting fundamentalthermodynamic linkage in DNA aptamers. Results are discussed in the context of thethermodynamic stability and structure of each of the aptamers examined.

  14. Electrochemical techniques for characterization of stem-loop probe and linear probe-based DNA sensors.

    Science.gov (United States)

    Lai, Rebecca Y; Walker, Bryce; Stormberg, Kent; Zaitouna, Anita J; Yang, Weiwei

    2013-12-15

    Here we present a summary of the sensor performance of the stem-loop probe (SLP) and linear probe (LP) electrochemical DNA sensors when interrogated using alternating current voltammetry (ACV), cyclic voltammetry (CV), and differential pulse voltammetry (DPV). Specifically, we identified one critical parameter for each voltammetric technique that can be adjusted for optimal sensor performance. Overall, the SLP sensor displayed good sensor performance (i.e., 60+% signal attenuation in the presence of the target) over a wider range of experimental conditions when compared to the LP sensor. When used with ACV, the optimal frequency range was found to be between 5 and 5000 Hz, larger than the 5-100 Hz range observed with the LP sensor. A similar trend was observed for the two sensors in CV; the LP sensor was operational only at scan rates between 30 and 100 V/s, whereas the SLP sensor performed well at scan rates between 1 and 1000 V/s. Unlike ACV and CV, DPV has demonstrated to be a more versatile sensor interrogation technique for this class of sensors. Despite the minor differences in total signal attenuation upon hybridization to the target DNA, both SLP and LP sensors performed optimally under most pulse widths used in this study. More importantly, when used with longer pulse widths, both sensors showed "signal-on" behavior, which is generally more desirable for sensor applications.

  15. Protocols for 16S rDNA Array Analyses of Microbial Communities by Sequence-Specific Labeling of DNA Probes

    Directory of Open Access Journals (Sweden)

    Knut Rudi

    2003-01-01

    Full Text Available Analyses of complex microbial communities are becoming increasingly important. Bottlenecks in these analyses, however, are the tools to actually describe the biodiversity. Novel protocols for DNA array-based analyses of microbial communities are presented. In these protocols, the specificity obtained by sequence-specific labeling of DNA probes is combined with the possibility of detecting several different probes simultaneously by DNA array hybridization. The gene encoding 16S ribosomal RNA was chosen as the target in these analyses. This gene contains both universally conserved regions and regions with relatively high variability. The universally conserved regions are used for PCR amplification primers, while the variable regions are used for the specific probes. Protocols are presented for DNA purification, probe construction, probe labeling, and DNA array hybridizations.

  16. DNA probes for papillomavirus strains readied for cervical cancer screening

    Energy Technology Data Exchange (ETDEWEB)

    Merz, B.

    1988-11-18

    New Papillomavirus tests are ready to come to the aid of the standard Papanicolauo test in screening for cervical cancer. The new tests, which detect the strains of human papillomavirus (HPV) most commonly associated with human cervical cancer, are designed to be used as an adjunct to rather than as a replacement for the Papanicolaou smears. Their developers say that they can be used to indicated a risk of developing cancer in women whose Papanicolaou smears indicate mild cervical dysplasia, and, eventually, to detect papillomavirus infection in normal Papanicolaou smears. The rationale for HPV testing is derived from a growing body of evidence that HPV is a major factor in the etiology of cervical cancer. Three HPV tests were described recently in Chicago at the Third International Conference on Human Papillomavirus and Squamous Cervical Cancer. Each relies on DNA probes to detect the presence of papillomavirus in cervical cells and/or to distinguish the strain of papillomavirus present.

  17. Mixed-Sequence Recognition of Double-Stranded DNA Using Enzymatically Stable Phosphorothioate Invader Probes

    Directory of Open Access Journals (Sweden)

    Brooke A. Anderson

    2015-07-01

    Full Text Available Development of probes that allow for sequence-unrestricted recognition of double-stranded DNA (dsDNA continues to attract much attention due to the prospect for molecular tools that enable detection, regulation, and manipulation of genes. We have recently introduced so-called Invader probes as alternatives to more established approaches such as triplex-forming oligonucleotides, peptide nucleic acids and polyamides. These short DNA duplexes are activated for dsDNA recognition by installment of +1 interstrand zippers of intercalator-functionalized nucleotides such as 2′-N-(pyren-1-ylmethyl-2′-N-methyl-2′-aminouridine and 2′-O-(pyren-1-ylmethyluridine, which results in violation of the nearest neighbor exclusion principle and duplex destabilization. The individual probes strands have high affinity toward complementary DNA strands, which generates the driving force for recognition of mixed-sequence dsDNA regions. In the present article, we characterize Invader probes that are based on phosphorothioate backbones (PS-DNA Invaders. The change from the regular phosphodiester backbone furnishes Invader probes that are much more stable to nucleolytic degradation, while displaying acceptable dsDNA-recognition efficiency. PS-DNA Invader probes therefore present themselves as interesting probes for dsDNA-targeting applications in cellular environments and living organisms.

  18. Detection of Babesia bigemina infection: use of a DNA probe - a review

    Directory of Open Access Journals (Sweden)

    Gerald M. Buening

    1992-01-01

    Full Text Available The development of a repetitive DNA probe for Babesia bigemina was reviewed. The original plasmid (p(Bbi16 contained an insert of B. bigemina DNA of approximately 6.3 kb. This probe has been evaluated for specificityand analytical sensitivity by dot hybridization with isolates from Mexico, the Caribbean region and Kenya. A partial restriction map has been constructed and insert fragments have been subcloned and utilized as specific DNA probes. A comparison of 32P labelled and non-radioactive DNA probes was presented. Non-radioctive detection systems that have been used include digoxigenin dUTP incorporation, and detection by colorimetric substrate methods. Derivatives from the original DNA probe have been utilized to detect B. bigemina infection in a experimentally inoculated cattle, b field exposed cattle, c infected Boophilus microplus ticks, and d the development of a PCR amplification system.

  19. Preparation of multi-locus DNA probe cocktail by liquid-phase reassociation.

    Science.gov (United States)

    Tamaki, Y; Fukuda, M; Kishida, T; Wang, W

    1995-08-01

    We developed a simple, rapid method for the preparation of a DNA-fingerprinting probe cocktail, and tested its usefulness in paternity testing. Exploiting the property of tandemly repetitive DNA segments to be rapidly renatured after heat denaturation, we enriched restriction fragments of a child's genomic DNA for minisatellites by liquid phase reassociation followed by capture with immobilized streptavidin. We amplified and simultaneously labeled the reassociation product by anchored PCR using a digoxigenin-labeling mixture. Using this probe cocktail, we were able to detect fingerprints of paternity case trios, and the results were corroborated by DNA fingerprinting with a commercially available probe as well as by conventional phenotyping. Our method enables one to prepare a fresh cocktail of probes from the DNA sample under study during the overnight electrophoresis and Southern transfer steps in DNA fingerprinting, and eliminates the need of having an expensive probe of limited shelf life. If one has a practical outlook on DNA fingerprinting and regard it as a preliminary test, one does not have to use a cloned DNA probe. The present study demonstrates that a multi-locus probe cocktail serves such a practical purpose.

  20. A comparative hybridization analysis of yeast DNA with Paramecium parafusin- and different phosphoglucomutase-specific probes.

    Science.gov (United States)

    Wyroba, E; Satir, B H

    2000-01-01

    Molecular probes designed for the parafusin (PFUS), the Paramecium exocytic-sensitive phosphoglycoprotein, gave distinct hybridization patterns in Saccharomyces cerevisiae genomic DNA when compared with different phosphoglucomutase specific probes. These include two probes identical to segments of yeast phosphoglucomutase (PGM) genes 1 and 2. Neither of the PGM probes revealed the 7.4 and 5.9 kb fragments in Bgl II-cut yeast DNA digest detected with the 1.6 kb cloned PFUS cDNA and oligonucleotide constructed to the PFUS region (insertion 3--I-3) not found in other species. PCR amplification with PFUS-specific primers generated yeast DNA-species of the predicted molecular size which hybridized to the I-3 probe. A search of the yeast genome database produced an unassigned nucleotide sequence that showed 55% identity to parafusin gene and 37% identity to PGM2 (the major isoform of yeast phosphoglucomutase) within the amplified region.

  1. AFM characterization of ss-DNA probes immobilization: a sequence effect on surface organization

    Energy Technology Data Exchange (ETDEWEB)

    Lallemand, D [Laboratoire d' Electronique, Optoelectronique et Microsystemes, Ecole Centrale de Lyon, 36 avenue Guy de Collongue, 69134 Ecully (France); Rouillat, M H [Laboratoire d' Electronique, Optoelectronique et Microsystemes, Ecole Centrale de Lyon, 36 avenue Guy de Collongue, 69134 Ecully (France); Dugas, V [BioTray, Ecole Normale Superieure de Lyon, 46 allee d' Italie, 69364 Lyon Cedex 07 (France); Chevolot, Y [Laboratoire d' Electronique, Optoelectronique et Microsystemes, Ecole Centrale de Lyon, 36 avenue Guy de Collongue, 69134 Ecully (France); Souteyrand, E [Laboratoire d' Electronique, Optoelectronique et Microsystemes, Ecole Centrale de Lyon, 36 avenue Guy de Collongue, 69134 Ecully (France); Phaner-Goutorbe, M [Laboratoire d' Electronique, Optoelectronique et Microsystemes, Ecole Centrale de Lyon, 36 avenue Guy de Collongue, 69134 Ecully (France)

    2007-03-15

    The biological sensitivity of a DNA chip depends on the molecular organization of the immobilized probe molecules, single stranded DNA (ss-DNA), on the substrate in terms of accessibility and non specific interactions between probes and substrate. In this article, Amplitude Modulation - Atomic Force Microscopy (AM-AFM) was used to characterize at a molecular scale, the morphological organization of different immobilized probes. In our system, three different ss-DNA were covalently grafted on a silicon substrate with the same deposit process. We studied the influence of probe length (25 bases, 12 bases) and sequence arrangement (two different 25 base oligoprobes) on the morphological organization. We showed that immobilized probes organize themselves in different structures depending on their sequence.

  2. Study on the Interaction of Mitomycin C with ct-DNA by Pd-Porphin Room Temperature Phosphorescence Probe

    Institute of Scientific and Technical Information of China (English)

    2001-01-01

    Anticancer drug Mitomycin C (MMC) quenches remarkably phosphorescence and reduces lifetime of phosphorescence probe, Pd-meso-tetrakis-(4-trimethylaminophenyl)porphin (Pd-TAPP), in the presence of calf thymus DNA. These results may be attributed to the site competition of MMC with the probe and electron transfer between MMC and probe. MMC also increases polarization degree of the probe by covalent drug-DNA or DNA-drug-DNA crosslinking.

  3. Applications of Fiberoptics-Based Nanosensors to Drug Discovery

    Science.gov (United States)

    Vo-Dinh, Tuan; Scaffidi, Jonathan; Gregas, Molly; Zhang, Yan; Seewaldt, Victoria

    2013-01-01

    Background Fiber-optic nanosensors are fabricated by heating and pulling optical fibers to yield sub-micron diameter tips, and have been used for in vitro analysis of individual living mammalian cells. Immobilization of bioreceptors (e.g., antibodies, peptides, DNA, etc) selective to target analyte molecules of interest provides molecular specificity. Excitation light can be launched into the fiber, and the resulting evanescent field at the tip of the nanofiber can be used to excite target molecules bound to the bioreceptor molecules. The fluorescence or surface-enhanced Raman scattering produced by the analyte molecules is detected using an ultra-sensitive photodetector. Objective This article provides an overview of the development and application of fiber-optic nanosensors for drug discovery. Conclusions The nanosensors provide minimally invasive tools to probe sub-cellular compartments inside single living cells for health effect studies (e.g., detection of benzopyrene adducts) and medical applications (e.g., monitoring of apoptosis in cells treated with anti-cancer drugs). PMID:23496274

  4. Surface density dependence of PCR amplicon hybridization on PNA/DNA probe layers

    DEFF Research Database (Denmark)

    Yao, Danfeng; Kim, Junyoung; Yu, Fang

    2005-01-01

    at an intermediate sodium concentration (approximately 100 mM). These effects were mainly ascribed to the electrostatic cross talk among the hybridized DNA molecules and the secondary structure of PCR amplicons. For the negatively charged DNA probes, the hybridization reaction was subjected additionally to the DNA...

  5. Non-Covalent Fluorescent Labeling of Hairpin DNA Probe Coupled with Hybridization Chain Reaction for Sensitive DNA Detection.

    Science.gov (United States)

    Song, Luna; Zhang, Yonghua; Li, Junling; Gao, Qiang; Qi, Honglan; Zhang, Chengxiao

    2016-04-01

    An enzyme-free signal amplification-based assay for DNA detection was developed using fluorescent hairpin DNA probes coupled with hybridization chain reaction (HCR). The hairpin DNAs were designed to contain abasic sites in the stem moiety. Non-covalent labeling of the hairpin DNAs was achieved when a fluorescent ligand was bound to the abasic sites through hydrogen bonding with the orphan cytosine present on the complementary strand, accompanied by quench of ligand fluorescence. As a result, the resultant probes, the complex formed between the hairpin DNA and ligand, showed almost no fluorescence. Upon hybridization with target DNA, the probe underwent a dehybridization of the stem moiety containing an abasic site. The release of ligand from the abasic site to the solution resulted in an effective fluorescent enhancement, which can be used as a signal. Compared with a sensing system without HCR, a 20-fold increase in the sensitivity was achieved using the sensing system with HCR. The fluorescent intensity of the sensing system increased with the increase in target DNA concentration from 0.5 nM to 100 nM. A single mismatched target ss-DNA could be effectively discriminated from complementary target DNA. Genotyping of a G/C single-nucleotide polymorphism of polymerase chain reaction (PCR) products was successfully demonstrated with the sensing system. Therefore, integrating HCR strategy with non-covalent labeling of fluorescent hairpin DNA probes provides a sensitive and cost-effective DNA assay.

  6. Iodination as a probe for small regions of disrupted secondary structure in double-stranded DNA

    DEFF Research Database (Denmark)

    Jensen, Kaj Frank; Nes, Ingolf F.; Wells, Robert D.

    1976-01-01

    Conditions were established where the thallium-catalyzed iodination of random coil DNA proceeded 100–200 times faster than for native DNA. This reaction was explored as a probe for localized regions of disrupted base pairs in duplex DNA. A heteroduplex was constructed between DNA fragments produced......, if they existed within an otherwise helical DNA fragment 789 base pairs long. Iodination studies were performed on superhelical SV40 DNA and on linear plac DNA. Analysis of the relative amount of iodine in restriction endonuclease fragments of these DNAs revealed the absence of localized single-stranded regions....

  7. A Microfluidic Microbeads Fluorescence Assay with Quantum Dots-Bead-DNA Probe.

    Science.gov (United States)

    Ankireddy, S R; Kim, Jongsung

    2016-03-01

    A microfluidic bead-based nucleic acid sensor for the detection of tumor causing N-Ras genes using quantum dots has been developed. Presently, quantum dots-bead-DNA probe based hybridization detection methods are often called as 'bead based assays' and their success is substantially influenced by the dispensing and manipulation capability of the microfluidic technology. This study reports the detection of N-Ras cancer gene by fluorescence quenching of quantum dots immobilized on the surface of polystyrene beads. A microfluidic chip was constructed in which the quantum dots-bead-DNA probes were packed in the channel. The target DNA flowed across the beads and hybridized with immobilized probe sequences. The target DNA can be detected by the fluorescence quenching of the quantum dots due to their transfer of emission energy to intercalation dye after DNA hybridization. The mutated gene also induces fluorescence quenching but with less degree than the perfectly complementary target DNA.

  8. HBx-DNA probe preparation and its application in study of hepatocarcinogenesis

    Science.gov (United States)

    Gao, Feng-Guang; Sun, Wen-Sheng; Cao, Ying-Lin; Zhang, Li-Ning; Song, Jing; Li, Hua-Fen; Yan, Shi-Kun

    1998-01-01

    AIM: To study the role of HBV especially HBx Open Reading Frame (ORF) in the development of hepatocellular carcinoma (HCC). METHODS: HBV 3.2 kb fragment was retrieved by digesting recombinant plasmid pBR322-2HBV with EcoR I, and HBx 0.59 kb fragments by digesting HBV-DNA with BamH I and Bgl II. These fragments were labelled with digoxigenin to get HBV-DNA and HBx-DNA probes. HBV-DNA was detected in HCC by dot blot and Southern blot hybridization with HBV-DNA probe, so the positive specimens in which HBV-DNA were integrated were selected. HBx-DNA was subsequently detected in the selected specimens with HBx-DNA probe. RESULTS: HBV-DNA was detected in 75% HCC, among which integrated type, integrated + free type covered 63.6% and 36.4%. There was no free type. HBx-DNA was detected in 90.5% specimens of integrated type. CONCLUSION: Hepatocarcinogenesis was highly related to HBV-DNA integration, and HBV-DNA mainly integrated into chromosome with incomplete virus DNA fragments among which HBx fragment was the predominant one. PMID:11819309

  9. Effect of salts, solvents and buffer on miRNA detection using DNA silver nanocluster (DNA/AgNCs) probes

    Science.gov (United States)

    Shah, Pratik; Cho, Seok Keun; Waaben Thulstrup, Peter; Bhang, Yong-Joo; Ahn, Jong Cheol; Choi, Suk Won; Rørvig-Lund, Andreas; Yang, Seong Wook

    2014-01-01

    MicroRNAs (miRNAs) are small regulatory RNAs (size ˜21 nt to ˜25 nt) which regulate a variety of important cellular events in plants, animals and single cell eukaryotes. Especially because of their use in diagnostics of human diseases, efforts have been directed towards the invention of a rapid, simple and sequence selective detection method for miRNAs. Recently, we reported an innovative method for the determination of miRNA levels using the red fluorescent properties of DNA/silver nanoclusters (DNA/AgNCs). Our method is based on monitoring the emission drop of a DNA/AgNCs probe in the presence of its specific target miRNA. Accordingly, the accuracy and efficiency of the method relies on the sensitivity of hybridization between the probe and target. To gain specific and robust hybridization between probe and target, we investigated a range of diverse salts, organic solvents, and buffer to optimize target sensing conditions. Under the newly adjusted conditions, the target sensitivity and the formation of emissive DNA/AgNCs probes were significantly improved. Also, fortification of the Tris-acetate buffer with inorganic salts or organic solvents improved the sensitivity of the DNA/AgNC probes. On the basis of these optimizations, the versatility of the DNA/AgNCs-based miRNA detection method can be expanded.

  10. Probing structural changes of self assembled i-motif DNA

    KAUST Repository

    Lee, Iljoon

    2015-01-01

    We report an i-motif structural probing system based on Thioflavin T (ThT) as a fluorescent sensor. This probe can discriminate the structural changes of RET and Rb i-motif sequences according to pH change. This journal is

  11. Immobilization-free electrochemical DNA detection with anthraquinone-labeled pyrrolidinyl peptide nucleic acid probe.

    Science.gov (United States)

    Kongpeth, Jutatip; Jampasa, Sakda; Chaumpluk, Piyasak; Chailapakul, Orawon; Vilaivan, Tirayut

    2016-01-01

    Electrochemical detection provides a simple, rapid, sensitive and inexpensive method for DNA detection. In traditional electrochemical DNA biosensors, the probe is immobilized onto the electrode. Hybridization with the DNA target causes a change in electrochemical signal, either from the intrinsic signal of the probe/target or through a label or a redox indicator. The major drawback of this approach is the requirement for probe immobilization in a controlled fashion. In this research, we take the advantage of different electrostatic properties between PNA and DNA to develop an immobilization-free approach for highly sequence-specific electrochemical DNA sensing on a screen-printed carbon electrode (SPCE) using a square-wave voltammetric (SWV) technique. Anthraquinone-labeled pyrrolidinyl peptide nucleic acid (AQ-PNA) was employed as a probe together with an SPCE that was modified with a positively-charged polymer (poly quaternized-(dimethylamino-ethyl)methacrylate, PQDMAEMA). The electrostatic attraction between the negatively-charged PNA-DNA duplex and the positively-charged modified SPCE attributes to the higher signal of PNA-DNA duplex than that of the electrostatically neutral PNA probe, resulting in a signal change. The calibration curve of this proposed method exhibited a linear range between 0.35 and 50 nM of DNA target with a limit of detection of 0.13 nM (3SD(blank)/Slope). The sub-nanomolar detection limit together with a small sample volume required (20 μL) allowed detection of DNA. With the high specificity of the pyrrolidinyl PNA probe used, excellent discrimination between complementary and various single-mismatched DNA targets was obtained. An application of this new platform for a sensitive and specific detection of isothermally-amplified shrimp's white spot syndrome virus (WSSV) DNA was successfully demonstrated.

  12. Buying Fiber-Optic Networks.

    Science.gov (United States)

    Fickes, Michael

    2003-01-01

    Describes consortia formed by college and university administrators to buy, manage, and maintain their own fiber-optic networks with the goals of cutting costs of leasing fiber-optic cable and planning for the future. Growth capacity is the real advantage of owning fiber-optic systems. (SLD)

  13. Fabrication of Unimolecular Double-stranded DNA Microarrays on Solid Surfaces for Probing DNA-Protein/Drug Interactions

    Directory of Open Access Journals (Sweden)

    Zuhong Lu

    2003-01-01

    Full Text Available We present a novel method for fabricating unimole cular double-stranded DNA microarrays on solid surfaces, which were used to probe sequence-specific DNA/protein interactions. For manufacturing the unimolecular double-stranded DNA microarrays, two kinds of special single-stranded oligonucleotides, constant oligonucleotide and target oligonucleotide, were chemically synthesized. The constant oligonucleotides with internal aminated dT were used to capture and immobilize the target oligonucleotides onto the solid surface, and also to provide a primer for later enzymatic extension reactions, while target oligonucleotides took the role of harbouring DNA-binding sites of DNA-binding proteins. The variant target oligonucleotides were annealed and ligated with the constant oligonucleotides to form the new unimolecular oligonucleotides for microspotting. The prepared unimolecular oligonucleotides were microspotted on aldehyde-derivatized glass slides to make partial-dsDNA microarrays. Finally, the partial-dsDNA microarrays were converted into a unimolecular complete-dsDNA microarray by a DNA polymerase extension reaction. The efficiency and accuracy of the polymerase synthesis were demonstrated by the fluorescent-labeled dUTP incorporation in the enzymatic extension reaction and the restriction endonuclease digestion of the fabricated unimolecular complete-dsDNA microarray. The accessibility and specificity of the sequence-specific DNA-binding proteins binding to the immobilized unimolecular dsDNA probes were demonstrated by the binding of Cy3 labeled NF-?B (p50·p50 to the unimolecular dsDNA microarray. This unimolecular dsDNA microarray provides a general technique for high-throughput DNA-protein or DNA-drugs interactions.

  14. Preparation of fluorescent DNA probe by solid-phase organic synthesis

    Directory of Open Access Journals (Sweden)

    2009-08-01

    Full Text Available Fluorescent DNA probe based on fluorescence resonance energy transfer (FRET was prepared by solid-phase organic synthesis when CdTe quantum dots (QDs were as energy donors and Au nanoparticles (AuNPs were as energy accepters. The poly(divinylbenzene core/poly(4-vinylpyridine shell microspheres, as solid-phase carriers, were prepared by seeds distillation-precipitation polymerization with 2,2′-azobisisobutyronitrile (AIBN as initiator in neat acetonitrile. The CdTe QDs and AuNPs were self-assembled on the surface of core/shell microspheres, and then the linkage of CdTe QDs with oligonucleotides (CdTe-DNA and AuNPs with complementary single-stranded DNA (Au-DNA was on the solid-phase carriers instead of in aqueous solution. The hybridization of complementary double stranded DNA (dsDNA bonded to the QDs and AuNPs (CdTe-dsDNA-Au determined the FRET distance of CdTe QDs and AuNPs. Compared with the fluorescence of CdTe-DNA, the fluorescence of CdTe-dsDNA-Au conjugates (DNA probes decreased extremely, which indicated that the FRET occurred between CdTe QDs and AuNPs. The probe system would have a certain degree recovery of fluorescence when the complementary single stranded DNA was introduced into this system, which showed that the distance between CdTe QDs and AuNPs was increased.

  15. Combining ligation reaction and capillary gel electrophoresis to obtain reliable long DNA probes.

    Science.gov (United States)

    García-Cañas, Virginia; Mondello, Monica; Cifuentes, Alejandro

    2011-05-01

    New DNA amplification methods are continuously developed for sensitive detection and quantification of specific DNA target sequences for, e.g. clinical, environmental or food applications. These new applications often require the use of long DNA oligonucleotides as probes for target sequences hybridization. Depending on the molecular technique, the length of DNA probes ranges from 40 to 450 nucleotides, solid-phase chemical synthesis being the strategy generally used for their production. However, the fidelity of chemical synthesis of DNA decreases for larger DNA probes. Defects in the oligonucleotide sequence result in the loss of hybridization efficiency, affecting the sensitivity and selectivity of the amplification method. In this work, an enzymatic procedure has been developed as an alternative to solid-phase chemical synthesis for the production of long oligonucleotides. The enzymatic procedure for probe production was based on ligation of short DNA sequences. Long DNA probes were obtained from smaller oligonucleotides together with a short sequence that acts as bridge stabilizing the molecular complex for DNA ligation. The ligation reactions were monitored by capillary gel electrophoresis with laser-induced fluorescence detection (CGE-LIF) using a bare fused-silica capillary. The capillary gel electrophoresis-LIF method demonstrated to be very useful and informative for the characterization of the ligation reaction, providing important information about the nature of some impurities, as well as for the fine optimization of the ligation conditions (i.e. ligation cycles, oligonucleotide and enzyme concentration). As a result, the yield and quality of the ligation product were highly improved. The in-lab prepared DNA probes were used in a novel multiplex ligation-dependent genome amplification (MLGA) method for the detection of genetically modified maize in samples. The great possibilities of the whole approach were demonstrated by the specific and sensitive

  16. Development of a specific DNA probe and PCR for the detection of Mycoplasma bovis.

    Science.gov (United States)

    Ghadersohi, A; Coelen, R J; Hirst, R G

    1997-05-01

    Mycoplasma bovis is responsible for several production diseases in cattle, including mastitis, arthritis, pneumonia, abortion and infertility. Current methodologies for detecting and identifying M. bovis are time consuming and difficult. Tests which rely on antigen or antibody detection have poor sensitivity and specificity. In this paper associated protocols for the development of a hybridization probe and PCR are described. A genomic library (SauIIIA digested) was prepared from M. bovis DNA (Colindale Reference Strain: NC10131:02) and cloned into pUC19. Colony hybridization, using a probe preparation made from purified M. bovis DNA, was used to identify colonies of interest. M. bovis DNA fragments were retrieved from recombinant plasmids by digestion with EcoRI and HindIII. This DNA was used to prepare randomly primed probes for dot blot hybridization analysis with immobilized DNA from M. bovis (two strains), M. dispar, M. agalactiae, M. bovigenitalium (two strains), M. ovipneumoniae, a Group 7 strain, M. arginini and bacteria belonging to different genera. Four probes were found to hybridize only with M. bovis and M. ovipneumoniae DNA, whereas one probe reacted with genomic DNA from only one of the two M. bovis strains. The level of sensitivity of the dot blot hybridization assay was 200 CFU (colony forming units)/mL. To enhance the sensitivity further, an M. bovis-specific PCR assay was developed. The primers were designed using sequences obtained from the probe DNA which discriminated M. bovis from all other Mycoplasma DNA tested. The minimum amount of target DNA that could be detected by the PCR assay was that isolated from 10-20 CFU/mL. The PCR assay was therefore 10 times more sensitive than dot blot hybridization.

  17. [Cu(phen)2](2+) acts as electrochemical indicator and anchor to immobilize probe DNA in electrochemical DNA biosensor.

    Science.gov (United States)

    Yang, Linlin; Li, Xiaoyu; Li, Xi; Yan, Songling; Ren, Yinna; Wang, Mengmeng; Liu, Peng; Dong, Yulin; Zhang, Chaocan

    2016-01-01

    We demonstrate a novel protocol for sensitive in situ label-free electrochemical detection of DNA hybridization based on copper complex ([Cu(phen)2](2+), where phen = 1,10-phenanthroline) and graphene (GR) modified glassy carbon electrode. Here, [Cu(phen)2](2+) acted advantageously as both the electrochemical indicator and the anchor for probe DNA immobilization via intercalative interactions between the partial double helix structure of probe DNA and the vertical aromatic groups of phen. GR provided large density of docking site for probe DNA immobilization and increased the electrical conductivity ability of the electrode. The modification procedure was monitored by electrochemical impedance spectroscopy (EIS). Square-wave voltammetry (SWV) was used to explore the hybridization events. Under the optimal conditions, the designed electrochemical DNA biosensor could effectively distinguish different mismatch degrees of complementary DNA from one-base mismatch to noncomplementary, indicating that the biosensor had high selectivity. It also exhibited a reasonable linear relationship. The oxidation peak currents of [Cu(phen)2](2+) were linear with the logarithm of the concentrations of complementary target DNA ranging from 1 × 10(-12) to 1 × 10(-6) M with a detection limit of 1.99 × 10(-13) M (signal/noise = 3). Moreover, the stability of the electrochemical DNA biosensor was also studied.

  18. Hairpin DNA probe based surface plasmon resonance biosensor used for the activity assay of E. coli DNA ligase.

    Science.gov (United States)

    Luan, Qingfen; Xue, Ying; Yao, Xin; Lu, Wu

    2010-02-01

    Using hairpin DNA probe self-structure change during DNA ligation process, a sensitive, label-free and simple method of E. coli DNA ligase assay via a home-built high-resolution surface plasmon resonance (SPR) instrument was developed. The DNA ligation process was monitored in real-time and the effects of single-base mutation on the DNA ligation process were investigated. Then an assay of E. coli DNA ligase was completed with a lower detection limit (0.6 nM), wider concentration range and better reproducibility. Moreover, the influence of Quinacrine on the activity of E. coli DNA ligase was also studied, which demonstrated that our method was useful for drug screening.

  19. LD-RTPCR:\tA NEW METHOD FOR LABELLING TRACE cDNA MICROARRAY PROBE

    Institute of Scientific and Technical Information of China (English)

    范保星; 孙敬芬; 梁好; 王升启; 周平坤; 吴德昌

    2002-01-01

    Objective: To explore the usefulness of long distance reverse transcript combining linear amplification (LD-RTPCR) in labeling slight trace probe used for cDNA microarray. Methods: Total RNA from BEP2D cells was extracted and labeled by two different methods, LD-RTPCR with Cy3-dCTP as fluorescent dye and traditionally used RNA reverse transcript (RT) with Cy5-dCTP as fluorescent dye. Then, the probes labeled by two methods were mixed equally and hybridized with the cDNA microarray. Results: Scan and analysis of the microarray showed that the two methods labeled probes had consistent results. Conclusion: LD-RTPCR was proved useful for labeling cDNA microarray probe, especially for limited RNA material.

  20. Data Mining Empowers the Generation of a Novel Class of Chromosome-specific DNA Probes

    Energy Technology Data Exchange (ETDEWEB)

    Zeng, Hui; Weier, Heinz-Ulrich G.; Kwan, Johnson; Wang, Mei; O' Brien, Benjamin

    2011-03-08

    Probes that allow accurate delineation of chromosome-specific DNA sequences in interphase or metaphase cell nuclei have become important clinical tools that deliver life-saving information about the gender or chromosomal make-up of a product of conception or the probability of an embryo to implant, as well as the definition of tumor-specific genetic signatures. Often such highly specific DNA probes are proprietary in nature and have been the result of extensive probe selection and optimization procedures. We describe a novel approach that eliminates costly and time consuming probe selection and testing by applying data mining and common bioinformatics tools. Similar to a rational drug design process in which drug-protein interactions are modeled in the computer, the rational probe design described here uses a set of criteria and publicly available bioinformatics software to select the desired probe molecules from libraries comprised of hundreds of thousands of probe molecules. Examples describe the selection of DNA probes for the human X and Y chromosomes, both with unprecedented performance, but in a similar fashion, this approach can be applied to other chromosomes or species.

  1. Label-free DNA hybridization detection by various spectroscopy methods using triphenylmethane dyes as a probe.

    Science.gov (United States)

    Tu, Jiaojiao; Cai, Changqun; Ma, Ying; Luo, Lin; Weng, Chao; Chen, Xiaoming

    2012-12-01

    A new assay is developed for direct detection of DNA hybridization using triphenylmethane dye as a probe. It is based on various spectroscopic methods including resonance light scattering (RLS), circular dichroism (CD), ultraviolet spectra and fluorescence spectra, as well as atomic force microscopy (AFM), six triphenylmethane dyes interact with double strand DNA (dsDNA) and single strand DNA (ssDNA) were investigated, respectively. The interaction results in amplified resonance light scattering signals and enables the detection of hybridization without the need for labeling DNA. Mechanism investigations have shown that groove binding occurs between dsDNA and these triphenylmethane dyes, which depends on G-C sequences of dsDNA and the molecular volumes of triphenylmethane dyes. Our present approaches display the advantages of simple and fast, accurate and reliable, and the artificial samples were determined with satisfactory results. Copyright © 2012 Elsevier B.V. All rights reserved.

  2. Probing the Conformational Distributions of Sub-Persistence Length DNA

    Energy Technology Data Exchange (ETDEWEB)

    Mastroianni, Alexander; Sivak, David; Geissler, Phillip; Alivisatos, Paul

    2009-06-08

    We have measured the bending elasticity of short double-stranded DNA (dsDNA) chains through small-angle X-ray scattering from solutions of dsDNA-linked dimers of gold nanoparticles. This method, which does not require exertion of external forces or binding to a substrate, reports on the equilibrium distribution of bending fluctuations, not just an average value (as in ensemble FRET) or an extreme value (as in cyclization), and in principle provides a more robust data set for assessing the suitability of theoretical models. Our experimental results for dsDNA comprising 42-94 basepairs (bp) are consistent with a simple worm-like chain model of dsDNA elasticity, whose behavior we have determined from Monte Carlo simulations that explicitly represent nanoparticles and their alkane tethers. A persistence length of 50 nm (150 bp) gave a favorable comparison, consistent with the results of single-molecule force-extension experiments on much longer dsDNA chains, but in contrast to recent suggestions of enhanced flexibility at these length scales.

  3. Effect of different concentration of HPV DNA probe immobilization for cervical cancer detection based IDE biosensor

    Science.gov (United States)

    Roshila, M. L.; Hashim, U.; Azizah, N.; Nadzirah, Sh.; Arshad, M. K. Md; Ruslinda, A. R.; Gopinath, Subash C. B.

    2017-03-01

    This paper principally delineates to the detection process of Human Papillomavirus (HPV) DNA test. HPV is an extremely common virus infection that infected to human by the progressions cell in the cervix cell. The types of HPV that give a most exceedingly awful infected with cervical cancer is 16 and 18 other than 31 and 45. The HPV DNA probe is immobilized with a different concentration to stabilize the sensitivity. A technique of rapid and sensitive for the HPV identification was proposed by coordinating basic DNA extraction with a quality of DNA. The extraction of the quality of DNA will make a proficiency of the discovery procedure. It will rely on the sequence of the capture probes and the way to support their attached. The fabrication, surface modification, immobilization and hybridization procedures are described by current-voltage (I-V) estimation by utilizing KEITHLEY 6487. This procedure will play out a decent affectability and selectivity of HPV discovery.

  4. Comet-FISH with rDNA probes for the analysis of mutagen-induced DNA damage in plant cells.

    Science.gov (United States)

    Kwasniewska, Jolanta; Grabowska, Marta; Kwasniewski, Miroslaw; Kolano, Bozena

    2012-06-01

    We used comet-fluorescence in situ hybridization (FISH) in the model plant species Crepis capillaris following exposure of seedlings to maleic hydrazide (MH). FISH with 5S and 25S rDNA probes was applied to comets obtained under alkaline conditions to establish whether these DNA regions were preferentially involved in comet tail formation. MH treatment induced significant fragmentation of nuclear DNA and of rDNA loci. A 24-h post-treatment recovery period allowed a partial reversibility of MH-induced damage on nuclear and rDNA regions. Analyses of FISH signals demonstrated that rDNA sequences were always involved in tail formation and that 5S rDNA was more frequently present in the tail than 25S rDNA, regardless of treatment. The involvement of 25S rDNA in nucleolus formation and differences in chromatin structure between the two loci may explain the different susceptibility of the 25S and 5S rDNA regions to migrate into the tail. This work is the first report on the application of FISH to comet preparations from plants to analyze the distribution and repair of DNA damage within specific genomic regions after mutagenic treatment. Moreover, our work suggests that comet-FISH in plants may be a useful tool for environmental monitoring assessment.

  5. Effect of structure on sensing performance of a target induced signaling probe shifting DNA-based (TISPS-DNA) sensor.

    Science.gov (United States)

    Yu, Xiang; Yu, Zhigang; Li, Fengqin; Xu, Yanmei; He, Xunjun; Xu, Lan; Shi, Wenbing; Zhang, Guiling; Yan, Hong

    2017-05-15

    A type of "signal on" displacement-based sensors named target induced signaling probe shifting DNA-based (TISPS-DNA) sensor were developed for a designated DNA detection. The signaling mechanism of the signaling probe (SP) shifting different from the classical conformation/flexibility change mode endows the sensor with high sensitivity. Through using thiolated or no thiolated capturing probe (CP), two 3-probe sensing structures, sensor-1 and sensor-2, were designed and constructed. The systematical comparing research results show that both sensors exhibit some similarities or big differences in sensing performance. On the one hand, the similarity in structures determines the similarity in some aspects of signaling mechanism, background signal, signal changing form, anti-fouling ability and versatility; on the other hand, the slight difference in structures also results in two opposite hybridization modes of gradual increasing resistance and gradual decreasing resistance which can affect the hybridization efficiency between the assistant probe (AP) and the SP, further producing some big differences in sensing performance, for example, apparently different signal enhancement (SE) change, point mutation discrimination ability and response speed. Under the optimized fabrication and detection conditions, both sensors feature high sensitivity for target DNAs with the detection limits of ∼10 fM for sensor-1 and ∼7 fM for sensor-2, respectively. Among many acquired sensing virtues, the sensor-1 shows a peculiar specificity adjustability which is also a highlight in this work. Copyright © 2017 Elsevier B.V. All rights reserved.

  6. Immobilization of human papillomavirus DNA probe for surface plasmon resonance imaging

    Science.gov (United States)

    Chong, Xinyuan; Ji, Yanhong; Ma, Suihua; Liu, Le; Liu, Zhiyi; Li, Yao; He, Yonghong; Guo, Jihua

    2009-08-01

    Human papillomavirus (HPV) is a kind of double-stranded DNA virus whose subspecies have diversity. Near 40 kinds of subspecies can invade reproductive organ and cause some high risk disease, such as cervical carcinoma. In order to detect the type of the subspecies of the HPV DNA, we used the parallel scan spectral surface plasmon resonance (SPR) imaging technique, which is a novel type of two- dimensional bio-sensing method based on surface plasmon resonance and is proposed in our previous work, to study the immobilization of the HPV DNA probes on the gold film. In the experiment, four kinds of the subspecies of the HPV DNA (HPV16, HPV18, HPV31, HPV58) probes are fixed on one gold film, and incubate in the constant temperature condition to get a HPV DNA probe microarray. We use the parallel scan spectral SPR imaging system to detect the reflective indices of the HPV DNA subspecies probes. The benefits of this new approach are high sensitive, label-free, strong specificity and high through-put.

  7. Coaxial atomic force microscope probes for dielectrophoresis of DNA under different buffer conditions

    Science.gov (United States)

    Tao, Yinglei; Kumar Wickramasinghe, H.

    2017-02-01

    We demonstrate a coaxial AFM nanoprobe device for dielectrophoretic (DEP) trapping of DNA molecules in Tris-EDTA (TE) and phosphate-buffered saline (PBS) buffers. The DEP properties of 20 nm polystyrene beads were studied with coaxial probes in media with different conductivities. Due to the special geometry of our DEP probe device, sufficiently high electric fields were generated at the probe end to focus DNA molecules with positive DEP. DEP trapping for both polystyrene beads and DNA molecules was quantitatively analyzed over the frequency range from 100 kHz to 50 MHz and compared with the Clausius-Mossotti theory. Finally, we discussed the negative effect of medium salinity during DEP trapping.

  8. Probing DNA with micro- and nanocapillaries and optical tweezers

    Energy Technology Data Exchange (ETDEWEB)

    Steinbock, L J; Otto, O; Skarstam, D R; Jahn, S; Chimerel, C; Gornall, J L; Keyser, U F, E-mail: ufk20@cam.ac.u [Cavendish Laboratory, University of Cambridge, J J Thomson Avenue, Cambridge CB3 0HE (United Kingdom)

    2010-11-17

    We combine for the first time optical tweezer experiments with the resistive pulse technique based on capillaries. Quartz glass capillaries are pulled into a conical shape with tip diameters as small as 27 nm. Here, we discuss the translocation of {lambda}-phage DNA which is driven by an electrophoretic force through the nanocapillary. The resulting change in ionic current indicates the folding state of single {lambda}-phage DNA molecules. Our flow cell design allows for the straightforward incorporation of optical tweezers. We show that a DNA molecule attached to an optically trapped colloid is pulled into a capillary by electrophoretic forces. The detected electrophoretic force is in good agreement with measurements in solid-state nanopores.

  9. Microarray long oligo probe designing for Escherichia coli: an in-silico DNA marker extraction.

    Science.gov (United States)

    Behzadi, Payam; Najafi, Ali; Behzadi, Elham; Ranjbar, Reza

    2016-01-01

    Urinary tract infections are predominant diseases which may be caused by different pathogenic microorganisms, particularly Escherichia coli (E.coli). DNA microarray technology is an accurate, rapid, sensitive, and specific diagnostic tool which may lead to definite diagnosis and treatment of several infectious diseases. DNA microarray is a multi-process method in which probe designing plays an important. Therefore, the authors of the present study have tried to design a range of effective and proper long oligo microarray probes for detection and identification of different strains of pathogenic E.coli and in particular, uropathogenic E.coli (UPEC). E.coli O26 H11 11368 uid41021 was selected as the standard strain for probe designing. This strain encompasses the largest nucleotide sequence and the most number of genes among other pathogenic strains of E.coli. For performing this in silico survey, NCBI database, GReview Server, PanSeq Server, Oligoanalyzer tool, and AlleleID 7.7 were used to design accurate, appropriate, effective, and flexible long oligo microarray probes. Moreover, the genome of E.coli and its closely related microorganisms were compared. In this study, 15 long oligo microarray probes were designed for detecting and identifying different strains of E.coli such as UPEC. These probes possessed the best physico-chemical characteristics. The functional and structural properties of the designed probes were recognized by practical tools and softwares. The use of reliable advanced technologies and methodologies for probe designing guarentees the high quality of microarray probes and makes DNA microarray technology more flexible and an effective diagnostic technique.

  10. An Integrated DNA Modified Dual-microelectrode Sensor Probe

    Institute of Scientific and Technical Information of China (English)

    Xiang Qin LIN; Li Ping LU; Xiao Hua JIANG

    2005-01-01

    A unique method for preparing a coaxial dual-microelectrode sensor by vaporizing the nano-thickness Au layer on the DNA modified carbon fiber micro-column electrode was illustrated.The dual-electrode showed particular merit for determination in biological systems.

  11. Probing the binding of coumarins and cyclothialidines to DNA gyrase

    DEFF Research Database (Denmark)

    Kampranis, S C; Gormley, N A; Tranter, R

    1999-01-01

    DNA gyrase is the target of a number of antibacterial agents, including the coumarins and the cyclothialidines. To extend our understanding of the mechanism of action of these compounds, we have examined the previously published crystal structures of the complexes between the 24 kDa fragment of Gyr...

  12. Increased detectability of somatic changes in the DNA from human tumours after probing with "synthetic" and "genome-derived" hypervariable multilocus probes

    DEFF Research Database (Denmark)

    Lagoda, P J; Seitz, G; Epplen, J T

    1989-01-01

    DNA fingerprinting with two minisatellite (33.15, M13) and two simple repeat probes [(GACA)4, (CAC)5/(GTG)s] was performed to screen for somatic changes in the DNA from various solid human tumours in comparison with constitutional DNA from the same patient. Loss of bands or changes in band...... intensities were observed. Together the probes 33.15 and (CAC)5/(GTG)5 detected deviating fingerprint patterns in 63% of the colorectal carcinomas investigated. In mammary and stomach carcinomas, only 1/11 and 2/11 tumours, respectively, showed differences with either of the three probes, 33.15, (GACA)4...

  13. Origin and molecular organization of supernumerary chromosomes of Prochilodus lineatus (characiformes, prochilodontidae) obtained by DNA probes.

    Science.gov (United States)

    Voltolin, Tatiana Aparecida; Laudicina, Alejandro; Senhorini, José Augusto; Bortolozzi, Jehud; Oliveira, Cláudio; Foresti, Fausto; Porto-Foresti, Fábio

    2010-12-01

    In Prochilodus lineatus B-chromosomes are visualized as reduced size extra elements identified as microchromosomes and are variable in morphology and number. We describe the specific total probe (B-chromosome probe) in P. lineatus obtained by chromosome microdissection and a whole genomic probe (genomic probe) from an individual without B-chromosome. The specific B-chromosome was scraped and processed to obtain DNA with amplification by DOP-PCR, and so did the genomic probe DNA. Fluorescence in situ hybridization using the B-chromosome probe labeled with dUTP-Tetramethyl-rhodamine and the genomic probe labeled with digoxigenin-FITC permitted to establish that in this species supernumerary chromosomes with varying number and morphology had different structure of chromatin when compared to that of the regular chromosomes or A complement, since only these extra elements were labeled in the metaphases. The present findings suggest that modifications in the chromatin structure of B-chromosomes to differentiate them from the A chromosomes could occur along their dispersion in the individuals of the population.

  14. Fiber-Optic Sensor Facility

    Data.gov (United States)

    Federal Laboratory Consortium — FUNCTION: Constructs and evaluates fiber-optic sensors for a variety of measurands. These measurands include acoustic, pressure, magnetic, and electric field as well...

  15. Study of concentration of HPV DNA probe immobilization for cervical cancer detection based IDE biosensor

    Science.gov (United States)

    Roshila, M. L.; Hashim, U.; Azizah, N.

    2016-07-01

    This paper mainly illustrates regarding the detection process of Human Papillomavirus (HPV) DNA probe. HPV is the most common virus that infected to human by a sexually transmitted virus. The most common high-risk HPV are 16 and 18. Interdigitated electrode (IDE) device used as based of Titanium Dioxide (TiO2) acts as inorganic surface, where by using APTES as a linker between inorganic surface and organic surface. A strategy of rapid and sensitive for the HPV detection was proposed by integrating simple DNA extraction with a gene of DNA. The extraction of the gene of DNA will make an efficiency of the detection process. It will depend on the sequence of the capture probes and the way to support their attached. The fabrication, surface modification, immobilization and hybridization processes are characterized by current voltage (I-V) measurement by using KEITHLEY 6487. This strategy will perform a good sensitivity of HPV detection.

  16. DNA aptamers as molecular probes for colorectal cancer study.

    Directory of Open Access Journals (Sweden)

    Kwame Sefah

    Full Text Available BACKGROUND: Understanding the molecular features of specific tumors can increase our knowledge about the mechanism(s underlying disease development and progression. This is particularly significant for colorectal cancer, which is a heterogeneous complex of diseases developed in a sequential manner through a multistep carcinogenic process. As such, it is likely that tumors with similar characteristics might originate in the same manner and have a similar molecular behavior. Therefore, specific mapping of the molecular features can be potentially useful for both tumor classification and the development of appropriate therapeutic regimens. However, this can only be accomplished by developing high-affinity molecular probes with the ability to recognize specific markers associated with different tumors. Aptamers can most easily meet this challenge based on their target diversity, flexible manipulation and ease of development. METHODOLOGY AND RESULTS: Using a method known as cell-based Systematic Evolution of Ligands by Exponential enrichment (cell-SELEX and colorectal cancer cultured cell lines DLD-1 and HCT 116, we selected a panel of target-specific aptamers. Binding studies by flow cytometry and confocal microscopy showed that these aptamers have high affinity and selectivity. Our data further show that these aptamers neither recognize normal colon cells (cultured and fresh, nor do they recognize most other cancer cell lines tested. CONCLUSION/SIGNIFICANCE: The selected aptamers can identify specific biomarkers associated with colorectal cancers. We believe that these probes could be further developed for early disease detection, as well as prognostic markers, of colorectal cancers.

  17. Probing DNA-Protein Interactions on Surfaces Using Spectral Self-interference Fluorescence Microscopy

    Science.gov (United States)

    Dogan, Mehmet; Droge, Peter; Swan, Anna K.; Unlu, Selim; Goldberg, Bennett B.

    2007-03-01

    We are probing the interactions between double-stranded DNA and integration host factor (IHF) proteins [1] on surfaces using Spectral Self-interference Fluorescence Microscopy (SSFM) [2].The probing technique utilizes the spectral fringes produced by interference of direct and reflected emission from fluorescent molecules. The modified spectrum provides a unique signature of the axial position of the fluorophores. Using the SSFM technique, we probe the average location of the fluorescent markers attached to the DNA molecules to study the conformational changes in double-stranded DNA tethered to SiO2 surfaces. In the presence of IHF, a DNA bending protein, we observe reduction in the vertical position of fluorescent molecules suggesting the formation of IHF-DNA complex and IHF-induced DNA bending. We also discuss the results with different IHF strains and different binding conditions. [1] Q. Bao et. al., Gene, Vol.343 pp.99-106 (2004) [2] L.A. Moiseev et. al., Journal of Applied Physics, Vol.96, pp. 5311-5315 (2004)

  18. Efficient in situ detection of mRNAs using the Chlorella virus DNA ligase for padlock probe ligation.

    Science.gov (United States)

    Schneider, Nils; Meier, Matthias

    2017-02-01

    Padlock probes are single-stranded DNA molecules that are circularized upon hybridization to their target sequence by a DNA ligase. In the following, the circulated padlock probes are amplified and detected with fluorescently labeled probes complementary to the amplification product. The hallmark of padlock probe assays is a high detection specificity gained by the ligation reaction. Concomitantly, the ligation reaction is the largest drawback for a quantitative in situ detection of mRNAs due to the low affinities of common DNA or RNA ligases to RNA-DNA duplex strands. Therefore, current protocols require that mRNAs be reverse transcribed to DNA before detection with padlock probes. Recently, it was found that the DNA ligase from Paramecium bursaria Chlorella virus 1 (PBCV-1) is able to efficiently ligate RNA-splinted DNA. Hence, we designed a padlock probe assay for direct in situ detection of mRNAs using the PBCV-1 DNA ligase. Experimental single-cell data were used to optimize and characterize the efficiency of mRNA detection with padlock probes. Our results demonstrate that the PBCV-1 DNA ligase overcomes the efficiency limitation of current protocols for direct in situ mRNA detection, making the PBCV-1 DNA ligase an attractive tool to simplify in situ ligation sequencing applications.

  19. Combination probes with intercalating anchors and proximal fluorophores for DNA and RNA detection.

    Science.gov (United States)

    Qiu, Jieqiong; Wilson, Adam; El-Sagheer, Afaf H; Brown, Tom

    2016-09-30

    A new class of modified oligonucleotides (combination probes) has been designed and synthesised for use in genetic analysis and RNA detection. Their chemical structure combines an intercalating anchor with a reporter fluorophore on the same thymine nucleobase. The intercalator (thiazole orange or benzothiazole orange) provides an anchor, which upon hybridisation of the probe to its target becomes fluorescent and simultaneously stabilizes the duplex. The anchor is able to communicate via FRET to a proximal reporter dye (e.g. ROX, HEX, ATTO647N, FAM) whose fluorescence signal can be monitored on a range of analytical devices. Direct excitation of the reporter dye provides an alternative signalling mechanism. In both signalling modes, fluorescence in the unhybridised probe is switched off by collisional quenching between adjacent intercalator and reporter dyes. Single nucleotide polymorphisms in DNA and RNA targets are identified by differences in the duplex melting temperature, and the use of short hybridization probes, made possible by the stabilisation provided by the intercalator, enhances mismatch discrimination. Unlike other fluorogenic probe systems, placing the fluorophore and quencher on the same nucleobase facilitates the design of short probes containing multiple modifications. The ability to detect both DNA and RNA sequences suggests applications in cellular imaging and diagnostics. © The Author(s) 2016. Published by Oxford University Press on behalf of Nucleic Acids Research.

  20. DNA-based digital tension probes reveal integrin forces during early cell adhesion

    Science.gov (United States)

    Zhang, Yun; Ge, Chenghao; Zhu, Cheng; Salaita, Khalid

    2014-10-01

    Mechanical stimuli profoundly alter cell fate, yet the mechanisms underlying mechanotransduction remain obscure because of a lack of methods for molecular force imaging. Here to address this need, we develop a new class of molecular tension probes that function as a switch to generate a 20- to 30-fold increase in fluorescence upon experiencing a threshold piconewton force. The probes employ immobilized DNA hairpins with tunable force response thresholds, ligands and fluorescence reporters. Quantitative imaging reveals that integrin tension is highly dynamic and increases with an increasing integrin density during adhesion formation. Mixtures of fluorophore-encoded probes show integrin mechanical preference for cyclized RGD over linear RGD peptides. Multiplexed probes with variable guanine-cytosine content within their hairpins reveal integrin preference for the more stable probes at the leading tip of growing adhesions near the cell edge. DNA-based tension probes are among the most sensitive optical force reporters to date, overcoming the force and spatial resolution limitations of traction force microscopy.

  1. Sub-diffusion and trapped dynamics of neutral and charged probes in DNA-protein coacervates

    Directory of Open Access Journals (Sweden)

    Najmul Arfin

    2013-11-01

    Full Text Available The physical mechanism leading to the formation of large intermolecular DNA-protein complexes has been studied. Our study aims to explain the occurrence of fast coacervation dynamics at the charge neutralization point, followed by the appearance of smaller complexes and slower coacervation dynamics as the complex experiences overcharging. Furthermore, the electrostatic potential and probe mobility was investigated to mimic the transport of DNA / DNA-protein complex in a DNA-protein complex coacervate medium [N. Arfin and H. B. Bohidar, J. Phys. Chem. B 116, 13192 (2012] by assigning neutral, negative, or positive charge to the probe particle. The mobility of the neutral probe was maximal at low matrix concentrations and showed random walk behavior, while its mobility ceased at the jamming concentration of c = 0.6, showing sub-diffusion and trapped dynamics. The positively charged probe showed sub-diffusive random walk followed by trapped dynamics, while the negatively charged probe showed trapping with occasional hopping dynamics at much lower concentrations. Sub-diffusion of the probe was observed in all cases under consideration, where the electrostatic interaction was used exclusively as the dominant force involved in the dynamics. For neutral and positive probes, the mean square displacement ⟨R2⟩ exhibits a scaling with time as ⟨R2⟩ ∼ tα, distinguishing random walk and trapped dynamics at α = 0.64 ± 0.04 at c = 0.12 and c = 0.6, respectively. In addition, the same scaling factors with the exponent β = 0.64 ± 0.04 can be used to distinguish random walk and trapped dynamics for the neutral and positive probes using the relation between the number of distinct sites visited by the probe, S(t, which follows the scaling, S(t ∼ tβ/ln (t. Our results established the occurrence of a hierarchy of diffusion dynamics experienced by a probe in a dense medium that is either charged or neutral.

  2. Sub-diffusion and trapped dynamics of neutral and charged probes in DNA-protein coacervates

    Science.gov (United States)

    Arfin, Najmul; Yadav, Avinash Chand; Bohidar, H. B.

    2013-11-01

    The physical mechanism leading to the formation of large intermolecular DNA-protein complexes has been studied. Our study aims to explain the occurrence of fast coacervation dynamics at the charge neutralization point, followed by the appearance of smaller complexes and slower coacervation dynamics as the complex experiences overcharging. Furthermore, the electrostatic potential and probe mobility was investigated to mimic the transport of DNA / DNA-protein complex in a DNA-protein complex coacervate medium [N. Arfin and H. B. Bohidar, J. Phys. Chem. B 116, 13192 (2012)] by assigning neutral, negative, or positive charge to the probe particle. The mobility of the neutral probe was maximal at low matrix concentrations and showed random walk behavior, while its mobility ceased at the jamming concentration of c = 0.6, showing sub-diffusion and trapped dynamics. The positively charged probe showed sub-diffusive random walk followed by trapped dynamics, while the negatively charged probe showed trapping with occasional hopping dynamics at much lower concentrations. Sub-diffusion of the probe was observed in all cases under consideration, where the electrostatic interaction was used exclusively as the dominant force involved in the dynamics. For neutral and positive probes, the mean square displacement ⟨R2⟩ exhibits a scaling with time as ⟨R2⟩ ˜ tα, distinguishing random walk and trapped dynamics at α = 0.64 ± 0.04 at c = 0.12 and c = 0.6, respectively. In addition, the same scaling factors with the exponent β = 0.64 ± 0.04 can be used to distinguish random walk and trapped dynamics for the neutral and positive probes using the relation between the number of distinct sites visited by the probe, S(t), which follows the scaling, S(t) ˜ tβ/ln (t). Our results established the occurrence of a hierarchy of diffusion dynamics experienced by a probe in a dense medium that is either charged or neutral.

  3. Sub-diffusion and trapped dynamics of neutral and charged probes in DNA-protein coacervates

    Energy Technology Data Exchange (ETDEWEB)

    Arfin, Najmul [Polymer and Biophysics Laboratory, School of Physical Sciences, Jawaharlal Nehru University, New Delhi-110067 (India); Yadav, Avinash Chand [Nonlinear Dynamics Laboratory, School of Physical Sciences, Jawaharlal Nehru University, New Delhi-110067 (India); Bohidar, H. B., E-mail: bohi0700@mail.jnu.ac.in [Polymer and Biophysics Laboratory, School of Physical Sciences, Jawaharlal Nehru University, New Delhi-110067 (India); Special Centre for Nanosciences, Jawaharlal Nehru University, New Delhi-110067 (India)

    2013-11-15

    The physical mechanism leading to the formation of large intermolecular DNA-protein complexes has been studied. Our study aims to explain the occurrence of fast coacervation dynamics at the charge neutralization point, followed by the appearance of smaller complexes and slower coacervation dynamics as the complex experiences overcharging. Furthermore, the electrostatic potential and probe mobility was investigated to mimic the transport of DNA / DNA-protein complex in a DNA-protein complex coacervate medium [N. Arfin and H. B. Bohidar, J. Phys. Chem. B 116, 13192 (2012)] by assigning neutral, negative, or positive charge to the probe particle. The mobility of the neutral probe was maximal at low matrix concentrations and showed random walk behavior, while its mobility ceased at the jamming concentration of c = 0.6, showing sub-diffusion and trapped dynamics. The positively charged probe showed sub-diffusive random walk followed by trapped dynamics, while the negatively charged probe showed trapping with occasional hopping dynamics at much lower concentrations. Sub-diffusion of the probe was observed in all cases under consideration, where the electrostatic interaction was used exclusively as the dominant force involved in the dynamics. For neutral and positive probes, the mean square displacement 〈R{sup 2}〉 exhibits a scaling with time as 〈R{sup 2}〉 ∼ t{sup α}, distinguishing random walk and trapped dynamics at α = 0.64 ± 0.04 at c = 0.12 and c = 0.6, respectively. In addition, the same scaling factors with the exponent β = 0.64 ± 0.04 can be used to distinguish random walk and trapped dynamics for the neutral and positive probes using the relation between the number of distinct sites visited by the probe, S(t), which follows the scaling, S(t) ∼ t{sup β}/ln (t). Our results established the occurrence of a hierarchy of diffusion dynamics experienced by a probe in a dense medium that is either charged or neutral.

  4. Enhancement of the immobi- lization and discrimination of DNA probe on a biosensor using gold nanoparticles

    Institute of Scientific and Technical Information of China (English)

    2001-01-01

    Nanoparticles have been widely utilized in biological research in recent years. 1,6-Hexanedithiol was used as the medium in this work to attach Au nanoparticles on thegold plates. We studied the immobilization and hybridization of DNA probes onto the resulted plates using the quartz crystal microbalance method. Approximately 3-5 times adsorption enhancement of HS-DNA on the plates was ob-tained under our experimental conditions. Using this DNA sensor, the sensitivity was promoted by approximately three times to 0.35 mg/mL.

  5. Cyanines as new fluorescent probes for DNA detection and two-photon excited bioimaging.

    Science.gov (United States)

    Feng, Xin Jiang; Wu, Po Lam; Bolze, Frédéric; Leung, Heidi W C; Li, King Fai; Mak, Nai Ki; Kwong, Daniel W J; Nicoud, Jean-François; Cheah, Kok Wai; Wong, Man Shing

    2010-05-21

    A series of cyanine fluorophores based on fused aromatics as an electron donor for DNA sensing and two-photon bioimaging were synthesized, among which the carbazole-based biscyanine exhibits high sensitivity and efficiency as a fluorescent light-up probe for dsDNA, which shows selective binding toward the AT-rich regions. The synergetic effect of the bischromophoric skeleton gives a several-fold enhancement in a two-photon absorption cross-section as well as a 25- to 100-fold enhancement in two-photon excited fluorescence upon dsDNA binding.

  6. Molecular detection of bacterial pathogens using microparticle enhanced double-stranded DNA probes.

    Science.gov (United States)

    Riahi, Reza; Mach, Kathleen E; Mohan, Ruchika; Liao, Joseph C; Wong, Pak Kin

    2011-08-15

    Rapid, specific, and sensitive detection of bacterial pathogens is essential toward clinical management of infectious diseases. Traditional approaches for pathogen detection, however, often require time-intensive bacterial culture and amplification procedures. Herein, a microparticle enhanced double-stranded DNA probe is demonstrated for rapid species-specific detection of bacterial 16S rRNA. In this molecular assay, the binding of the target sequence to the fluorophore conjugated probe thermodynamically displaces the quencher probe and allows the fluorophore to fluoresce. By incorporation of streptavidin-coated microparticles to localize the biotinylated probes, the sensitivity of the assay can be improved by 3 orders of magnitude. The limit of detection of the assay is as few as eight bacteria without target amplification and is highly specific against other common pathogens. Its applicability toward clinical diagnostics is demonstrated by directly identifying bacterial pathogens in urine samples from patients with urinary tract infections.

  7. Probing the mechanics of the complete DNA transcription cycle in real-time using optical tweezers.

    Science.gov (United States)

    Baumann, Christoph G; Cross, Stephen J

    2011-01-01

    RNA polymerase (RNAP) is a DNA-dependent motor protein that links ribonucleotide polymerization to force generation and DNA translocation through its active site, i.e., mechanical work. Single-molecule studies using optical tweezers have allowed researchers to probe the load-dependent ribonucleotide incorporation rate and processivity of both single-subunit viral and multisubunit prokaryotic and eukaryotic RNAPs engaged in transcription elongation. A single-molecule method is described here, which allows the complete transcription cycle (i.e., promoter binding, initiation, elongation and termination) to be followed in real-time using dual-trap optical tweezers and a unique "three-bead" geometry. This single-molecule transcription assay can be used to probe the mechanics of both stationary and moving RNAP-DNA complexes engaged in different stages of transcription.

  8. DNA binding activity of Anabaena sensory rhodopsin transducer probed by fluorescence correlation spectroscopy.

    Science.gov (United States)

    Kim, Sung Hyun; Kim, So Young; Jung, Kwang-Hwan; Kim, Doseok

    2015-01-01

    Anabaena sensory rhodopsin transducer (ASRT) is believed to be a major player in the photo-signal transduction cascade, which is triggered by Anabaena sensory rhodopsin. Here, we characterized DNA binding activity of ASRT probed by using fluorescence correlation spectroscopy. We observed clear decrease of diffusion coefficient of DNA upon binding of ASRT. The dissociation constant, K(D), of ASRT to 20 bp-long DNA fragments lied in micro-molar range and varied moderately with DNA sequence. Our results suggest that ASRT may interact with several different regions of DNA with different binding affinity for global regulation of several genes that need to be activated depending on the light illumination.

  9. Multiplex DNA assay based on nanoparticle probes by single particle inductively coupled plasma mass spectrometry.

    Science.gov (United States)

    Zhang, Shixi; Han, Guojun; Xing, Zhi; Zhang, Sichun; Zhang, Xinrong

    2014-04-01

    A multiplex DNA assay based on nanoparticle (NP) tags detection utilizing single particle mode inductively coupled plasma mass spectrometry (SP-ICP-MS) as ultrasensitive readout has been demonstrated in the article. Three DNA targets associated with clinical diseases (HIV, HAV, and HBV) down to 1 pM were detected by DNA probes labeled with AuNPs, AgNPs, and PtNPs via DNA sandwich assay. Single nucleotide polymorphisms in genes can also be effectively discriminated. Since our method is unaffected by the sample matrix, it is well-suited for diagnostic applications. Moreover, with the high sensitivity of SP-ICP-MS and the variety of NPs detectable by SP-ICP-MS, high-throughput DNA assay could be achieved without signal amplification or chain reaction amplification.

  10. Multi-Probe Based Artificial DNA Encoding and Matching Classifier for Hyperspectral Remote Sensing Imagery

    Directory of Open Access Journals (Sweden)

    Ke Wu

    2016-08-01

    Full Text Available In recent years, a novel matching classification strategy inspired by the artificial deoxyribonucleic acid (DNA technology has been proposed for hyperspectral remote sensing imagery. Such a method can describe brightness and shape information of a spectrum by encoding the spectral curve into a DNA strand, providing a more comprehensive way for spectral similarity comparison. However, it suffers from two problems: data volume is amplified when all of the bands participate in the encoding procedure and full-band comparison degrades the importance of bands carrying key information. In this paper, a new multi-probe based artificial DNA encoding and matching (MADEM method is proposed. In this method, spectral signatures are first transformed into DNA code words with a spectral feature encoding operation. After that, multiple probes for interesting classes are extracted to represent the specific fragments of DNA strands. During the course of spectral matching, the different probes are compared to obtain the similarity of different types of land covers. By computing the absolute vector distance (AVD between different probes of an unclassified spectrum and the typical DNA code words from the database, the class property of each pixel is set as the minimum distance class. The main benefit of this strategy is that the risk of redundant bands can be deeply reduced and critical spectral discrepancies can be enlarged. Two hyperspectral image datasets were tested. Comparing with the other classification methods, the overall accuracy can be improved from 1.22% to 10.09% and 1.19% to 15.87%, respectively. Furthermore, the kappa coefficient can be improved from 2.05% to 15.29% and 1.35% to 19.59%, respectively. This demonstrated that the proposed algorithm outperformed other traditional classification methods.

  11. The isothermal amplification detection of double-stranded DNA based on a double-stranded fluorescence probe.

    Science.gov (United States)

    Shi, Chao; Shang, Fanjin; Pan, Mei; Liu, Sen; Ma, Cuiping

    2016-06-15

    Here we have developed a novel method of isothermal amplification detection of double-stranded DNA (dsDNA) based on double-stranded fluorescence probe (ds-probe). Target dsDNA repeatedly generated single-stranded DNA (ssDNA) with polymerase and nicking enzyme. The ds-probe as a primer hybridized with ssDNA and extended to its 5'-end. The displaced ssDNA served as a new detection target to initiate above-described reaction. Meanwhile, the extended ds-probe could dynamically dissociate from ssDNA and self-hybridize, converting into a turn-back structure to initiate another amplification reaction. In particular, the ds-probe played a key role in the entire experimental process, which not only was as a primer but also produced the fluorescent signal by an extension and displacement reaction. Our method could detect the pBluescript II KS(+) plasmid with a detection limit of 2.3 amol, and it was also verified to exhibit a high specificity, even one-base mismatch. Overall, it was a true isothermal dsDNA detection strategy with a strongly anti-jamming capacity and one-pot, only requiring one ds-probe, which greatly reduced the cost and the probability of contamination. With its advantages, the approach of dsDNA detection will offer a promising tool in the field of point-of-care testing (POCT).

  12. Feasibility of Ultra-Thin Fiber-Optic Dosimeters for Radiotherapy Dosimetry

    Directory of Open Access Journals (Sweden)

    Bongsoo Lee

    2015-11-01

    Full Text Available In this study, prototype ultra-thin fiber-optic dosimeters were fabricated using organic scintillators, wavelength shifting fibers, and plastic optical fibers. The sensor probes of the ultra-thin fiber-optic dosimeters consisted of very thin organic scintillators with thicknesses of 100, 150 and 200 μm. These types of sensors cannot only be used to measure skin or surface doses but also provide depth dose measurements with high spatial resolution. With the ultra-thin fiber-optic dosimeters, surface doses for gamma rays generated from a Co-60 therapy machine were measured. Additionally, percentage depth doses in the build-up regions were obtained by using the ultra-thin fiber-optic dosimeters, and the results were compared with those of external beam therapy films and a conventional fiber-optic dosimeter.

  13. Differential diagnosis of Taenia saginata and Taenia solium infections: from DNA probes to polymerase chain reaction.

    Science.gov (United States)

    González, Luis Miguel; Montero, Estrella; Sciutto, Edda; Harrison, Leslie J S; Parkhouse, R Michael E; Garate, Teresa

    2002-04-01

    The objective of this work was the rapid and easy differential diagnosis of Taenia saginata and T. solium. First, a T. saginata size-selected genomic deoxyribonucleic acid (gDNA) library was constructed in the vector lambda gt10 using the 2-4 kb fraction from the parasite DNA digested with EcoR1, under 'star' conditions. After differential screening of the library and hybridization analysis with DNA from T. saginata, T. solium, T. taeniaeformis, T. crassiceps, and Echinococcus granulosus (bovine, porcine, and human), 2 recombinant phages were selected. They were designated HDP1 and HDP2. HDP1 reacted specifically with T. saginata DNA, and HDP2 recognized DNA from both T. saginata and T. solium. The 2 DNA probes were then sequenced and further characterized. HDP1 was a repetitive sequence with a 53 bp monomeric unit repeated 24 times in direct tandem along the 1272 bp fragment, while the 3954 bp HDP2 was not a repetitive sequence. Using the sequencing data, oligonucleotides were designed and used in a polymerase chain reaction (PCR). The 2 selected oligonucleotides from probe HDP1 (PTs4F1 and PTs4R1) specifically amplified gDNA from T. saginata, but not T. solium or other related cestodes, with a sensitivity of < 10 pg of T. saginata gDNA, about the quantity of DNA in one taeniid egg. The 3 oligonucleotides selected from the HDP2 sequence (PTs7S35F1, PTs7S35F2, and PTs7S35R1) allowed the differential amplification of gDNA from T. saginata, T. solium and E. granulosus in a multiplex PCR, again with a sensitivity of < 10 pg. These diagnostic tools have immediate application in the differential diagnosis of T. solium and T. saginata in humans and in the diagnosis of dubious cysts in the slaughterhouse. We also hope to apply them to epidemiological surveys of, for example, soil and water in endemic areas.

  14. Chemiluminescence-imaging detection of DNA on a solid-phase membrane by using a peroxidase-labeled macromolecular probe.

    Science.gov (United States)

    Azam, Md Golam; Yamasuji, Mutsumi; Krawczyk, Tomasz; Shibata, Takayuki; Kabashima, Tsutomu; Kai, Masaaki

    2015-07-01

    We have developed a novel method for sensitive chemiluminescence (CL)-imaging detection of DNA by using a macromolecular probe synthesized by attaching multiple molecules of horseradish peroxidase (HRP) and biotin in dextran backbone. The probe formed a macromolecular assembly by binding to streptavidin which specifically recognized biotinylated complementary DNA, which was hybridized to a target DNA on a solid-phase membrane. This methodology was applied to CL-imaging detection of a synthetic telomere DNA (TTAGGG)10 and human telomere DNA by using the CL probe comprising of dextranT2000 (MW=ca. 2000kDa) bonded to approximately 42 molecules of HRP and 210 molecules of biotin. The human telomere DNA in a small number of buccal mucous cells (ca. 70 cell numbers) of cheek tissue was quantitatively determined by the proposed CL detection method that afforded approximately 10 times higher sensitivity than that of the conventional CL method using commercially available HRP-avidin probe.

  15. Single and multiple molecular beacon probes for DNA hybridization studies on a silica glass surface

    Science.gov (United States)

    Fang, Xiaohong; Liu, Xiaojing; Tan, Weihong

    1999-05-01

    Surface immobilizable molecular beacons have been developed for DNA hybridization studies on a silica glass plate. Molecular beacons are a new class of oligonucleotide probes that have a loop-and-stem structure with a fluorophore and a quencher attached to the two ends of the stem. They only emit intense fluorescence when hybridize to their target molecules. This provides an excellent selectivity for the detection of DNA molecules. We have designed biotinylated molecular beacons which can be immobilized onto a solid surface. The molecular beacon is synthesized using DABCYL as the quencher and an optical stable dye, tetramethylrhodamine, as the fluorophore. Mass spectrometry is used to confirm the synthesized molecular beacon. The molecular beacons have been immobilized onto a silica surface through biotin-avidin binding. The surface immobilized molecular beacons have been used for the detection of target DNA with subnanomolar analytical sensitivity. have also immobilized two different molecular beacons on a silica surface in spatially resolved microscopic regions. The hybridization study of these two different molecular beacon probes has shown excellent selectivity for their target sequences. The newly designed molecular beacons are intended for DNA molecular interaction studies at an interface and for the development of ultrasensitive DNA sensors for a variety of applications including disease diagnosis, disease mechanism studies, new drug development, and in the investigation of molecular interactions between DNA molecules and other interesting biomolecules.

  16. [Fluorescence in situ hybridization with DNA probes derived from individual chromosomes and chromosome regions].

    Science.gov (United States)

    Bogomolov, A G; Karamysheva, T V; Rubtsov, N B

    2014-01-01

    A significant part of the eukaryotic genomes consists of repetitive DNA, which can form large clusters or distributed along euchromatic chromosome regions. Repeats located in chromosomal regions make a problem in analysis and identification of the chromosomal material with fluorescence in situ hybridization (FISH). In most cases, the identification of chromosome regions using FISH requires detection of the signal produced with unique sequences. The feasibility, advantages and disadvantages of traditional methods of suppression of repetitive DNA hybridization, methods of repeats-free probe construction and methods of chromosome-specific DNA sequences visualization using image processing of multicolor FISH results are considered in the paper. The efficiency of different techniques for DNA probe generation, different FISH protocols, and image processing of obtained microscopic images depends on the genomic size and structure of analyzing species. This problem was discussed and different approaches were considered for the analysis of the species with very large genome, rare species and species which specimens are too small in size to obtain the amount of genomic and Cot-1 DNA required for suppression of repetitive DNA hybridization.

  17. Study on the toxic interactions of Ni{sup 2+} with DNA using neutral red dye as a fluorescence probe

    Energy Technology Data Exchange (ETDEWEB)

    Wang Li [Shandong Key Laboratory of Water Pollution Control and Resource Reuse, School of Environmental Science and Engineering, Shandong University (China); CRC for Environment and Health, Shandong Province, 27 Shanda South Road, Jinan 250100 (China); Liu Rutao, E-mail: rutaoliu@sdu.edu.c [Shandong Key Laboratory of Water Pollution Control and Resource Reuse, School of Environmental Science and Engineering, Shandong University (China); CRC for Environment and Health, Shandong Province, 27 Shanda South Road, Jinan 250100 (China); Teng Yue [Shandong Key Laboratory of Water Pollution Control and Resource Reuse, School of Environmental Science and Engineering, Shandong University (China); CRC for Environment and Health, Shandong Province, 27 Shanda South Road, Jinan 250100 (China)

    2011-04-15

    The interaction between Ni{sup 2+} and calf thymus DNA (ctDNA) was investigated in simulated physiological buffer (pH 7.4) using the Neutral Red (NR) dye as a spectral probe by UV-vis absorption and fluorescence spectroscopy, as well as CD spectra. The experimental results showed that the conformational changes in DNA helix induced by Ni{sup 2+} are the reason for the fluorescence quenching of the DNA-NR system. From the experimental results, conclusion can be drawn that Ni{sup 2+} can cause structural changes of ctDNA and bind with DNA by electrostatic interaction. At the same time, the paper proved that conformation changes of DNA can also lead to the fluorescence decrease of DNA-probe systems. - Research Highlights: The formation of new non-fluorescence complex or competing binding sites between small molecules and probe are usually the reason of fluorescence quenching in DNA-probe systems. This study proved that conformation changes of DNA induced by Ni{sup 2+} can also lead to the fluorescence decrease of DNA-probe systems.

  18. Probing DNA-DNA Interactions with a Combination of Quadruple-Trap Optical Tweezers and Microfluidics.

    Science.gov (United States)

    Brouwer, Ineke; King, Graeme A; Heller, Iddo; Biebricher, Andreas S; Peterman, Erwin J G; Wuite, Gijs J L

    2017-01-01

    DNA metabolism and DNA compaction in vivo involve frequent interactions of remote DNA segments, mediated by proteins. In order to gain insight into such interactions, quadruple-trap optical tweezers have been developed. This technique provides an unprecedented degree of control through the ability to independently manipulate two DNA molecules in three dimensions. In this way, discrete regions of different DNA molecules can be brought into contact with one another, with a well-defined spatial configuration. At the same time, the tension and extension of the DNA molecules can be monitored. Furthermore, combining quadruple-trap optical tweezers with microfluidics makes fast buffer exchange possible, which is important for in situ generation of the dual DNA-protein constructs needed for these kinds of experiments. In this way, processes such as protein-mediated inter-DNA bridging can be studied with unprecedented control. This chapter provides a step-by-step description of how to perform a dual DNA manipulation experiment using combined quadruple-trap optical tweezers and microfluidics.

  19. A sensitive DNA biosensor based on a facile sulfamide coupling reaction for capture probe immobilization

    Energy Technology Data Exchange (ETDEWEB)

    Wang, Qingxiang, E-mail: axiang236@126.com [Department of Chemistry and Environment Science, Zhangzhou Normal University, Zhangzhou 363000 (China); Ding, Yingtao; Gao, Feng [Department of Chemistry and Environment Science, Zhangzhou Normal University, Zhangzhou 363000 (China); Jiang, Shulian [Zhangzhou Product Quality Supervision and Inspection Institute, Zhangzhou 363000 (China); Zhang, Bin; Ni, Jiancong; Gao, Fei [Department of Chemistry and Environment Science, Zhangzhou Normal University, Zhangzhou 363000 (China)

    2013-07-25

    Graphical abstract: A novel DNA biosensor was fabricated through a facile sulfamide coupling reaction between probe DNA and the sulfonic dye of 1-amino-2-naphthol-4-sulfonic acid that electrodeposited on a glassy carbon electrode. -- Highlights: •A versatile sulfonic dye of ANS was electrodeposited on a GCE. •A DNA biosensor was fabricated based on a facile sulfamide coupling reaction. •High probe DNA density of 3.18 × 10{sup 13} strands cm{sup −2} was determined. •A wide linear range and a low detection limit were obtained. -- Abstract: A novel DNA biosensor was fabricated through a facile sulfamide coupling reaction. First, the versatile sulfonic dye molecule of 1-amino-2-naphthol-4-sulfonate (AN-SO{sub 3}{sup −}) was electrodeposited on the surface of a glassy carbon electrode (GCE) to form a steady and ordered AN-SO{sub 3}{sup −} layer. Then the amino-terminated capture probe was covalently grafted to the surface of SO{sub 3}{sup −}-AN deposited GCE through the sulfamide coupling reaction between the amino groups in the probe DNA and the sulfonic groups in the AN-SO{sub 3}{sup −}. The step-by-step modification process was characterized by electrochemistry and attenuated total reflectance Fourier transform infrared (ATR-FTIR) spectroscopy. Using Ru(NH{sub 3}){sub 6}{sup 3+} as probe, the probe density and the hybridization efficiency of the biosensor were determined to be 3.18 × 10{sup 13} strands cm{sup −2} and 86.5%, respectively. The hybridization performance of the biosensor was examined by differential pulse voltammetry using Co(phen){sub 3}{sup 3+/2+} (phen = 1,10-phenanthroline) as the indicator. The selectivity experiments showed that the biosensor presented distinguishable response after hybridization with the three-base mismatched, non-complementary and complementary sequences. Under the optimal conditions, the oxidation peak currents of Co(phen){sub 3}{sup 3+/2+} increased linearly with the logarithm values of the concentration

  20. Honey, I Shrunk the DNA : DNA Length as a Probe for Nucleic-Acid Enzyme Activity

    NARCIS (Netherlands)

    Oijen, Antoine M. van

    2007-01-01

    The replication, recombination, and repair of DNA are processes essential for the maintenance of genomic information and require the activity of numerous enzymes that catalyze the polymerization or digestion of DNA. This review will discuss how differences in elastic properties between single- and d

  1. Honey, I Shrunk the DNA : DNA Length as a Probe for Nucleic-Acid Enzyme Activity

    NARCIS (Netherlands)

    Oijen, Antoine M. van

    2007-01-01

    The replication, recombination, and repair of DNA are processes essential for the maintenance of genomic information and require the activity of numerous enzymes that catalyze the polymerization or digestion of DNA. This review will discuss how differences in elastic properties between single- and d

  2. Improving Probe Immobilization for Label-Free Capacitive Detection of DNA Hybridization on Microfabricated Gold Electrodes

    Directory of Open Access Journals (Sweden)

    Sandro Carrara

    2008-02-01

    Full Text Available Alternative approaches to labeled optical detection for DNA arrays are actively investigated for low-cost point-of-care applications. In this domain, label-free capacitive detection is one of the most intensely studied techniques. It is based on the idea to detect the Helmholtz ion layer displacements when molecular recognition occurs at the electrodes/solution interface. The sensing layer is usually prepared by using thiols terminated DNA single-strength oligonucleotide probes on top of the sensor electrodes. However, published data shows evident time drift, which greatly complicates signal conditioning and processing and ultimately increases the uncertainty in DNA recognition sensing. The aim of this work is to show that newly developed ethylene-glycol functionalized alkanethiols greatly reduce time drift, thereby significantly improving capacitance based label-free detection of DNA.

  3. DNA detection using water-soluble conjugated polymers and peptide nucleic acid probes

    Science.gov (United States)

    Gaylord, Brent S.; Heeger, Alan J.; Bazan, Guillermo C.

    2002-08-01

    The light-harvesting properties of cationic conjugated polymers are used to sensitize the emission of a dye on a specific peptide nucleic acid (PNA) sequence for the purpose of homogeneous, "real-time" DNA detection. Signal transduction is controlled by hybridization of the neutral PNA probe and the negative DNA target. Electrostatic interactions bring the hybrid complex and cationic polymer within distances required for Förster energy transfer. Conjugated polymer excitation provides fluorescein emission >25 times higher than that obtained by exciting the dye, allowing detection of target DNA at concentrations of 10 pM with a standard fluorometer. A simple and highly sensitive assay with optical amplification that uses the improved hybridization behavior of PNA/DNA complexes is thus demonstrated.

  4. DNA probes for distinguishing Psychodopygus wellcomei from Psychodopygus complexus (Diptera: Psychodidae

    Directory of Open Access Journals (Sweden)

    P. D. Ready

    1991-03-01

    Full Text Available Genomic DNA fragments from males of Psychodopygus wellcomei were isolated and shown to be useful as sensitive diagnostic probles for positively separting individuals of this species from those of Ps. complexus. These two members of the Ps. squamiventris series are found sympatrically in foci of cutaneous leishmaniasis in the hill forests of southern Pará State. Of the two species, only Ps. welcomei is thought to be an important vector of Leishmania braziliensis sensu stricto, buth this is based on circumstantial evidence because of the difficulties of identifying female sandflies wothin the series. The diagnostic probes were isolated from a library of Ps. wellcomei built by ligationg short fragments of Sau 3A-resistricted, genomic DNA into the plasmid vector PUC 18. Differential screening of 1316 library clones with total genomic DNA of Ps. Wellcomei and Ps. complexus identified 5 recombinants, with cross-hybridizing inserts of repetitive DNA, that showed strong specificity for Ps. wellcomei. As little as 0.4% of the DNA extracted from an individual sandfly (=ca. 0.5 namograms was specifically detected. The diagnostic probes were used to identify as Ps. wellcomei a wild-caught female sandfly found infected with L. braziliensis s.s., providing only the second positive association between these two species.

  5. Nanoparticle probes and mid-infrared chemical imaging for DNA microarray detection.

    Science.gov (United States)

    Mossoba, Magdi M; Al-Khaldi, Sufian F; Schoen, Brianna; Yakes, Betsy Jean

    2010-11-01

    To date most mid-infrared spectroscopic studies have been limited, due to lack of sensitivity, to the structural characterization of a single oligonucleotide probe immobilized over the entire surface of a gold-coated slide or other infrared substrate. By contrast, widely used and commercially available glass slides and a microarray spotter that prints approximately 120-μm-diameter DNA spots were employed in the present work. To our knowledge, mid-infrared chemical imaging (IRCI) in the external reflection mode has been applied in the present study for the first time to the detection of nanostructure-based DNA microarrays spotted on glass slides. Alkyl amine-modified oligonucleotide probes were immobilized on glass slides that had been prefunctionalized with succinimidyl ester groups. This molecular fluorophore-free method entailed the binding of gold-nanoparticle-streptavidin conjugates to biotinylated DNA targets. Hybridization was visualized by the silver enhancement of gold nanoparticles. The adlayer of silver, selectively bound only to hybridized spots in a microarray, formed the external reflective infrared substrate that was necessary for the detection of DNA hybridization by IRCI in the present proof-of-concept study. IRCI made it possible to discriminate between diffuse and specular external reflection modes. The promising qualitative results are presented herein, and the implications for quantitative determination of DNA microarrays are discussed.

  6. Synthesis of RNA probes by the direct in vitro transcription of PCR-generated DNA templates.

    Science.gov (United States)

    Urrutia, R; McNiven, M A; Kachar, B

    1993-05-01

    We describe a novel method for the generation of RNA probes based on the direct in vitro transcription of DNA templates amplified by polymerase chain reaction (PCR) using primers with sequence hybrids between the target gene and those of the T7 and T3 RNA polymerases promoters. This method circumvents the need for cloning and allows rapid generation of strand-specific RNA molecules that can be used for the identification of genes in hybridization experiments. We have successfully applied this method to the identification of DNA sequences by Southern blot analysis and library screening.

  7. Fluorescent Probes Detecting the Phagocytic Phase of Apoptosis: Enzyme-Substrate Complexes of Topoisomerase and DNA

    Directory of Open Access Journals (Sweden)

    Candace L. Minchew

    2011-06-01

    Full Text Available In apoptosis, the initial self-driven suicide phase generates cellular corpses which are digested in the phagolysosomes of professional and amateur phagocytes during the subsequent waste-management phase. This ensures the complete elimination of the genetic material which often contains pathological, viral or cancerous DNA sequences. Although the phagocytic phase is critical for the efficient execution of apoptosis, there are currently few methods specifically adapted for its detailed visualization in the fixed tissue section format. To resolve this we developed new fluorescent probes for in situ research. The probes selectively visualize active phagocytic cells of any lineage (professional, amateur phagocytes or surrounding tissue cells which engulf and digest apoptotic cell DNA. These fluorescent probes are the covalently-bound enzyme-DNA intermediates produced in a topoisomerase reaction with specific “starting” oligonucleotides. They detect a specific marker of DNase II cleavage activity, which occurs exclusively in phagolysosomes of the cells that engulfed apoptotic nuclei. The probes provide snap-shot images of the digestion process occurring in cellular organelles responsible for the actual execution of phagocytic degradation of apoptotic cell corpses. We applied the probes for visualization of the phagocytic reaction in tissue sections of normal thymus and in several human lymphomas. We also discuss the nature, stability and properties of DNase II-type breaks as a marker of phagocytic activity. This development provides a useful fluorescent tool for studies of pathologies where clearance of dying cells is essential, such as cancers, inflammation, infection and auto-immune disorders.

  8. A highly polymorphic locus in human DNA revealed by cosmid-derived probes.

    OpenAIRE

    Litt, M.; White, R. L.

    1985-01-01

    Human gene mapping would be greatly facilitated if marker loci with sufficient heterozygosity were generally available. As a source of such markers, we have used cosmids from a human genomic library. We have developed a rapid method for screening random cosmids to identify those that are homologous to genomic regions especially rich in restriction fragment length polymorphisms. This method allows whole cosmids to be used as probes against Southern transfers of genomic DNA; regions of cosmid p...

  9. DNA quantification via ICP-MS using lanthanide-labeled probes and ligation-mediated amplification.

    Science.gov (United States)

    Brückner, Kathrin; Schwarz, Kathleen; Beck, Sebastian; Linscheid, Michael W

    2014-01-07

    The combination of lanthanide-tagged oligonucleotide probes with inductively coupled plasma mass spectrometry (ICP-MS) as the detection technique is a novel labeling and analysis strategy for heterogeneous nucleic acid quantification assays. We describe a hybridization assay based on biotin-streptavidin affinity using lanthanide-labeled reporter probes and biotinylated capture probes. For the basic sandwich type assay, performed in streptavidin-coated microtitration wells, the limit of detection (LOD) was 7.2 fmol of DNA target, corresponding to a final concentration of 6 pM terbium-labeled probes detectable by ICP-MS after elution from the solid support. To improve the sensitivity and sequence specificity of the approach, it was combined with established molecular biological techniques, i.e., elution with a restriction endonuclease and signal and target amplification by the ligase detection reaction (LDR) and ligase chain reaction (LCR), respectively. Initial experiments showed that the enzymes facilitated the discrimination of single-base mismatches within the recognition or ligation site. Furthermore, LCR as a target amplification step resulted in a 6000-fold increase of sensitivity, and finally an LOD of 2.6 amol was achieved with an artificial double-stranded DNA target.

  10. Characterization of an In Vivo Z-DNA Detection Probe Based on a Cell Nucleus Accumulating Intrabody.

    Science.gov (United States)

    Gulis, Galina; Silva, Izabel Cristina Rodrigues; Sousa, Herdson Renney; Sousa, Isabel Garcia; Bezerra, Maryani Andressa Gomes; Quilici, Luana Salgado; Maranhao, Andrea Queiroz; Brigido, Marcelo Macedo

    2016-09-01

    Left-handed Z-DNA is a physiologically unstable DNA conformation, and its existence in vivo can be attributed to localized torsional distress. Despite evidence for the existence of Z-DNA in vivo, its precise role in the control of gene expression is not fully understood. Here, an in vivo probe based on an anti-Z-DNA intrabody is proposed for native Z-DNA detection. The probe was used for chromatin immunoprecipitation of potential Z-DNA-forming sequences in the human genome. One of the isolated putative Z-DNA-forming sequences was cloned upstream of a reporter gene expression cassette under control of the CMV promoter. The reporter gene encoded an antibody fragment fused to GFP. Transient co-transfection of this vector along with the Z-probe coding vector improved reporter gene expression. This improvement was demonstrated by measuring reporter gene mRNA and protein levels and the amount of fluorescence in co-transfected CHO-K1 cells. These results suggest that the presence of the anti-Z-DNA intrabody can interfere with a Z-DNA-containing reporter gene expression. Therefore, this in vivo probe for the detection of Z-DNA could be used for global correlation of Z-DNA-forming sequences and gene expression regulation.

  11. Accuracy of the Clinical Diagnosis of Vaginitis Compared to a DNA Probe Laboratory Standard

    Science.gov (United States)

    Lowe, Nancy K.; Neal, Jeremy L.; Ryan-Wenger, Nancy A.

    2009-01-01

    Objective To estimate the accuracy of the clinical diagnosis of the three most common causes of acute vulvovaginal symptoms (bacterial vaginosis, candidiasis vaginitis, and trichomoniasis vaginalis) using a traditional, standardized clinical diagnostic protocol compared to a DNA probe laboratory standard. Methods This prospective clinical comparative study had a sample of 535 active duty United States military women presenting with vulovaginal symptoms. Clinical diagnoses were made by research staff using a standardized protocol of history, physical examination including pelvic examination, determination of vaginal pH, vaginal fluid amines test, and wet-prep microscopy. Vaginal fluid samples were obtained for DNA analysis. The research clinicians were blinded to the DNA results. Results The participants described a presenting symptom of abnormal discharge (50%), itching/irritation (33%), malodor (10%), burning (4%), or others such as vulvar pain and vaginal discomfort. According to laboratory standard, there were 225 cases (42%) of bacterial vaginosis 76 cases (14%) of candidiasis vaginitis, 8 cases (1.5%) of trichomoniasis vaginalis, 87 cases of mixed infections (16%), and 139 negative cases (26%). For each single infection, the clinical diagnosis had a sensitivity and specificity of 80.8% and 70.0% for bacterial vaginosis; 83.8% and 84.8% for candidiasis vaginitis; and 84.6% and 99.6% for trichomoniasis vaginalis when compared to the DNA probe standard. Conclusion Compared to a DNA probe standard, clinical diagnosis is 81-85% sensitive and 70- 99% specific for bacterial vaginosis, candida vaginitis, and trichomoniasis. Even under research conditions that provided clinicians with sufficient time and materials to conduct a thorough and standardized clinical evaluation, the diagnosis and therefore, subsequent treatment of these common vaginal problems remains difficult. PMID:19104364

  12. BaitFisher: A Software Package for Multispecies Target DNA Enrichment Probe Design.

    Science.gov (United States)

    Mayer, Christoph; Sann, Manuela; Donath, Alexander; Meixner, Martin; Podsiadlowski, Lars; Peters, Ralph S; Petersen, Malte; Meusemann, Karen; Liere, Karsten; Wägele, Johann-Wolfgang; Misof, Bernhard; Bleidorn, Christoph; Ohl, Michael; Niehuis, Oliver

    2016-07-01

    Target DNA enrichment combined with high-throughput sequencing technologies is a powerful approach to probing a large number of loci in genomes of interest. However, software algorithms that explicitly consider nucleotide sequence information of target loci in multiple reference species for optimizing design of target enrichment baits to be applicable across a wide range of species have not been developed. Here we present an algorithm that infers target DNA enrichment baits from multiple nucleotide sequence alignments. By applying clustering methods and the combinatorial 1-center sequence optimization to bait design, we are able to minimize the total number of baits required to efficiently probe target loci in multiple species. Consequently, more loci can be probed across species with a given number of baits. Using transcript sequences of 24 apoid wasps (Hymenoptera: Crabronidae, Sphecidae) from the 1KITE project and the gene models of Nasonia vitripennis, we inferred 57,650, 120-bp-long baits for capturing 378 coding sequence sections of 282 genes in apoid wasps. Illumina reduced-representation library sequencing confirmed successful enrichment of the target DNA when applying these baits to DNA of various apoid wasps. The designed baits furthermore enriched a major fraction of the target DNA in distantly related Hymenoptera, such as Formicidae and Chalcidoidea, highlighting the baits' broad taxonomic applicability. The availability of baits with broad taxonomic applicability is of major interest in numerous disciplines, ranging from phylogenetics to biodiversity monitoring. We implemented our new approach in a software package, called BaitFisher, which is open source and freely available at https://github.com/cmayer/BaitFisher-package.git. © The Author 2016. Published by Oxford University Press on behalf of the Society for Molecular Biology and Evolution. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

  13. DETECTION OF BRUGIA MALAYI INFECTED MOSQUITOES WITH SPECIES SPECIFIC DNA PROBE pBm 15, IN RIAU, INDONESIA

    Directory of Open Access Journals (Sweden)

    L. Kurniawan

    2012-09-01

    Full Text Available A species specific DNA probe (pBm15 was used in a field area where 2 filarial infections coexist: B.malayi in man and B.pahangi in cats. In our laboratory in Jakarta, this DNA probe proved to be sensitive enough to detect 500 ng DNA. One to two infective larvae of B.malayi could be detected with ease. This DNA probe did not react with infective larvae of wuchereria bancrofti, B.pahangi, and Dirofilaria spp. Non specific binding caused by undefined mosquito components was overcome with proteinase K and chitinase treatment. This additional step, made it possible for whole body mosquitoes to be squashed directly onto nitrocellulose paper. A comparative study of experimental infections of laboratory bred mosquitoes infected with B.malayi, showed no difference in infection rate between the group examined by dissection or by DNA probing. Mosquitoes which are vectors in Riau were collected and fed on microfilaremic patients of Riau. The set of mosquitoes were tested in parallel with mosquitoes infected with B.pahangi from cats. All fed mosquitoes were tested after 10-12 days. Only mosquitoes infected with B.malayi reacted in the assay. This study shows a success in applying the DNA probe technique in Jakarta. Further application in the field should be encouraged, with some modification of the DNA probing technique, for cheaper and easier implementation.

  14. Chromosomal DNA probes for the identification of asaccharolytic anaerobic pigmented bacterial rods from the oral cavity of cats.

    Science.gov (United States)

    Love, D N; Bailey, G D; Bastin, D

    1992-06-01

    A dot-blot hybridisation assay using isolated high molecular weight DNA as whole chromosomal probes of the cat pigmented asaccharolytic Bacteroides/Porphyromonas species was used against both purified high molecular weight DNA and DNA released on membranes from whole cells for the identification of B. salivosus and for its differentiation from the other anaerobic species isolated from normal and diseased mouths of cats and horses. 32P-labelled probes were compared with digoxigenin (DIG)-labelled probes (Boehringer-Mannheim). The whole chromosomal probes were specific--differentiating B. salivosus from a variety of species (including members of the genera Bacteroides, Fusobacterium, Eubacterium, and Prevotella) found in normal and abnormal mouths of cats and horses. Likewise, asaccharolytic black pigmented Group 2 strains were distinguishable from all strains tested. However, cat strains of P. gingivalis which show 68-76% DNA-DNA homology with human strain P. gingivalis ATCC 33277T, were not distinguishable from each other using either 32P-labelled or DIG-labelled probes. The minimum amount of pure Bacteroides DNA which could be detected by the 32P-labelled probe was 100-300 pg, while the amount of pure DNA detected by the DIG system was 1-3 mg after room temperature colour development for 1 h and 100-300 pg after 6 h colour development.

  15. A biostatistical study into the efficiency of individualism using nonisotopic chemiluminescent-enhanced NICE multilocus DNA probes.

    Science.gov (United States)

    Hau, P P; Watt, E H; Hau, C M

    1997-10-01

    The efficiency of individualisation using nonisotopic chemiluminescent- enhanced probes (NICE) was investigated by analysing DNA fingerprints obtained from 190 unrelated Caucasians. Novel analysis of the scoring procedure enabled us to include the comparison of 585 pairs of samples for each of two probes. When the results of NICE probes 33.6 and 33.15 were combined, the mean percentage band share between two unrelated individuals was 16.8% and the mean number of bands identified in an individual DNA fingerprint was 54.8. Results were compared with those obtained using isotopically labelled probes and suggest that the two labelling systems gave similar efficiencies for differentiating between individuals. Analysis of DNA fingerprints from 37 family trios (mother, child and father groups) gave a mutation rate of 0.10% when using NICE probes. The two labelling systems compared were equally efficient in establishing family relationships.

  16. Modeling Hybridization Kinetics of Gene Probes in a DNA Biochip Using FEMLAB

    Directory of Open Access Journals (Sweden)

    Ahsan Munir

    2017-05-01

    Full Text Available Microfluidic DNA biochips capable of detecting specific DNA sequences are useful in medical diagnostics, drug discovery, food safety monitoring and agriculture. They are used as miniaturized platforms for analysis of nucleic acids-based biomarkers. Binding kinetics between immobilized single stranded DNA on the surface and its complementary strand present in the sample are of interest. To achieve optimal sensitivity with minimum sample size and rapid hybridization, ability to predict the kinetics of hybridization based on the thermodynamic characteristics of the probe is crucial. In this study, a computer aided numerical model for the design and optimization of a flow-through biochip was developed using a finite element technique packaged software tool (FEMLAB; package included in COMSOL Multiphysics to simulate the transport of DNA through a microfluidic chamber to the reaction surface. The model accounts for fluid flow, convection and diffusion in the channel and on the reaction surface. Concentration, association rate constant, dissociation rate constant, recirculation flow rate, and temperature were key parameters affecting the rate of hybridization. The model predicted the kinetic profile and signal intensities of eighteen 20-mer probes targeting vancomycin resistance genes (VRGs. Predicted signal intensities and hybridization kinetics strongly correlated with experimental data in the biochip (R2 = 0.8131.

  17. Real-time electrochemical PCR with a DNA intercalating redox probe.

    Science.gov (United States)

    Deféver, Thibaut; Druet, Michel; Evrard, David; Marchal, Damien; Limoges, Benoit

    2011-03-01

    The proof-of-principle of a nonoptical real-time PCR method based on the electrochemical monitoring of a DNA intercalating redox probe that becomes considerably less easily electrochemically detectable once intercalated to the amplified double-stranded DNA is demonstrated. This has been made possible thanks to the finding of a redox intercalator that (i) strongly and specifically binds to the amplified double-stranded DNA, (ii) does not significantly inhibit PCR, (iii) is chemically stable under PCR cycling, and (iv) is sensitively detected by square wave voltammetry during PCR cycling. Among the different DNA intercalating redox probes that we have investigated, namely, methylene blue, Os[(bpy)(2)phen](2+), Os[(bpy)(2)DPPZ](2+), Os[(4,4'-dimethyl-bpy)(2)DPPZ](2+) and Os[(4,4'-diamino-bpy)(2)DPPZ](2+) (with bpy = 2,2'-bipyridine, phen = phenanthroline, and DPPZ = dipyrido[3,2-a:2',3'-c]phenazine), the one and only compound with which it has been possible to demonstrate the proof-of-concept is the Os[(bpy)(2)DPPZ](2+). In terms of analytical performances, the methodology described here compares well with optical-based real-time PCRs, offering finally the same advantages than the popular and routinely used SYBR Green-based real-time fluorescent PCR, but with the additional incomes of being potentially much cheaper and easier to integrate in a hand-held miniaturized device.

  18. A novel DNA tetrahedron-hairpin probe for in situ"off-on" fluorescence imaging of intracellular telomerase activity.

    Science.gov (United States)

    Feng, Qiu-Mei; Zhu, Meng-Jiao; Zhang, Ting-Ting; Xu, Jing-Juan; Chen, Hong-Yuan

    2016-04-21

    A novel three-dimensionally structured DNA probe is reported to realize in situ"off-on" imaging of intracellular telomerase activity. The probe consists of a DNA tetrahedron and a hairpin DNA on one of the vertices of the DNA tetrahedron. It is composed of four modified DNA segments: S1-Au nanoparticle (NP) inserting a telomerase strand primer (TSP) and S2-S4, three Cy5 dye modified DNA segments. Fluorescence of Cy5 at three vertices of the DNA tetrahedron is quenched by the Au NP at the other vertex due to the effective fluorescence resonance energy transfer (FRET) ("off" state). When the probe meets telomerase, the hairpin structure changes to rod-like through complementary hybridization with the telomerase-triggered stem elongation product, resulting in a large distance between the Au NP and Cy5 and the recovery of Cy5 fluorescence ("on" state). The molar ratio of 3 : 1 between the reporter (Cy5) and the target related TSP makes the probe show high sensitivity and recovery efficiency of Cy5 in the presence of telomerase extracted from HeLa cells. Given the functional and compact nanostructure, the mechanically stable and noncytotoxic nature of the DNA tetrahedron, this FRET-based probe provides more opportunities for biosensing, molecular imaging and drug delivery.

  19. Chromosomal DNA probes for the identification of Bacteroides tectum and Bacteroides fragilis from the oral cavity of cats.

    Science.gov (United States)

    Love, D N; Bailey, G D

    1993-01-01

    A dot-blot hybridisation assay using high molecular weight DNA as whole chromosomal probes was used to differentiate Bacteroides tectum from Bacteroides fragilis. 32P-labelled probes were compared with digoxigenin (DIG)-labelled probes. The whole chromosomal probes were specific--differentiating B. tectum from B. fragilis and both from a variety of other species (including other members of the genera Bacteroides, Fusobacterium, Eubacterium, and Prevotella) found in normal and abnormal mouths of cats and horses. However, even at very high stringencies, B. tectum homology groups I, II and III were not distinguishable from one another using either 32P-labelled or DIG-labelled probes. Thus, DIG-labelled whole chromosome probes directed against cellular DNA released directly onto nitrocellulose membranes is considered a useful method for diagnostic veterinary laboratories wishing to identify B. tectum and distinguish it from B. fragilis and other oral anaerobic flora of cats.

  20. Application of restriction display PCR technique in the preparation of cDNA microarray probes

    Institute of Scientific and Technical Information of China (English)

    Zhao-Hui Sun; Wen-Li Ma; Bao Zhang; Yi-Fei Peng; Wen-Ling Zheng

    2005-01-01

    AIM: To develop a simplified and efficient method for the preparation of hepatitis C virus (HCV) cDNA microarray probes.METHODS: With the technique of restriction display PCR (RD-PCR), restriction enzyme Sau3A I was chosen to digest the full-length HCV cDNAs. The products were classified and re-amplified by RD-PCR. We separated the differential genes by polyacrylamide gel electrophoresis and silver staining. Single bands cut out from the polyacrylamide gel were isolated. The third-round PCR was performed using the single bands as PCR template.The RD-PCR fragments were purified and cloned into the pMD18-T vector. The recombinant plasmids were extracted from positive clones, and the target gene fragments were sequenced. The cDNA microarray was prepared by spotting RD-PCR products to the surface of amino-modified glass slides using a robot. We validated the detection of microarray by hybridization and sequence analysis.RESULTS: A total of 24 different cDNA fragments ranging from 200 to 800 bp were isolated and sequenced,which were the specific gene fragments of HCV. These fragments could be further used as probes in microarray preparation. The diagnostic capability of the microarray was evaluated after the washing and scanning steps. The results of hybridization and sequence analysis showed that the specificity, sensitivity, accuracy, reproducibility,and linearity in detecting HCV RNA were satisfactory.CONCLUSION: The RD-PCR technique is of great value in obtaining a large number of size-comparable gene probes, which provides a speedy protocol in generating probes for the preparation of microarrays. Microarray prepared as such could be further optimized and applied in the clinical diagnosis of HCV.

  1. Cloning chromosome specific genes by reciprocal probing of arrayed cDNA and cosmid libraries

    Energy Technology Data Exchange (ETDEWEB)

    Yazdani, A.; Lee, C.C.; Wehnert, M. [Baylor College of Medicine, Houston, TX (United States)] [and others

    1994-09-01

    A human gene map will greatly facilitate the association of genes to single locus diseases and provide candidates for genes involved in complex genetic traits. Given the estimated 100,000 human genes an integrated strategy with a high throughput approach for isolation and mapping of expressed sequences is needed to create such a gene map. We have developed an approach that allows high throughput gene isolation and mapping using arrayed genomic and cDNA lambda libraries. Reciprocal probing of the arrayed genomic and cDNA cosmic libraries can rapidly establish cDNA-cosmid associations. Fluorescence in situ hybridization (FISH) chromosomal mapping and expressed sequence tag/sequence tag site (EST/STS) primers generated from DNA sequence of PCR-based mapping using somatic hybrid cell line mapping panels were utilized to characterize further the hybridization-based cDNA cosmid association. We have applied this approach to chromosome 17 using a placental cDNA library and have identified a total of 30 genes out of which 11 are novel. Furthermore seven cDNAs were mapped to 17q21 in this study, providing novel candidate genes for BRCA-1 gene for early onset breast cancer. The results of our study clearly show that an integration of an expression map into physical and genetic maps can provide candidate genes for human diseases that have been mapped to specific regions. This approach combined with the current physical mapping efforts could efficiently provide a detailed human gene map.

  2. Photocatalytic probing of DNA sequence by using TiO{sub 2}/dopamine-DNA triads

    Energy Technology Data Exchange (ETDEWEB)

    Liu Jianqin [Center for Nanoscale Materials, Argonne National Laboratory, 9700 South Cass Avenue, Argonne, IL 60439 (United States); Garza, Linda de la [Chemistry Division, Argonne National Laboratory, 9700 South Cass Avenue, Argonne, IL 60439 (United States); Zhang Ligang; Dimitrijevic, Nada M. [Center for Nanoscale Materials, Argonne National Laboratory, 9700 South Cass Avenue, Argonne, IL 60439 (United States); Zuo Xiaobing; Tiede, David M. [Chemistry Division, Argonne National Laboratory, 9700 South Cass Avenue, Argonne, IL 60439 (United States); Rajh, Tijana [Center for Nanoscale Materials, Argonne National Laboratory, 9700 South Cass Avenue, Argonne, IL 60439 (United States); Chemistry Division, Argonne National Laboratory, 9700 South Cass Avenue, Argonne, IL 60439 (United States)], E-mail: rajh@anl.gov

    2007-10-15

    A method to control charge transfer reaction in DNA using hybrid nanometer-sized TiO{sub 2} nanoparticles was developed. In this system extended charge separation reflects the sequence of DNA and was measured using metallic silver deposition or by photocurrent response. Light-induced extended charge separation in these systems was found to be dependent on the DNA-bridge length and sequence. The yield of photocatalytic deposition of silver was studied in systems having GG accepting sites imbedded in AT runs at varying distances from the TiO{sub 2} nanoparticle surface. Weak distance dependence of charge separation indicative of a hole hopping through mediating adenine (A) sites was found. The quantum yield of silver deposition in the system having a GG accepting site placed 8.5 A from the nanoparticle surface was found to be {phi} = 0.70 (70%) and {phi} = 0.56 (56%) for (A){sub n} and (AT){sub n/2} bridge, respectively. Hole injection to GG trapping sites as far as 70 A from a nanoparticle surface in the absence of G hopping sites was measured. Introduction of G hopping sites increased the efficiency of hole injection. The efficiency of photocatalytic deposition of metallic silver was found to be sensitive to the presence of a single nucleobase mismatch in the DNA sequence.

  3. DNA binding studies of hematoxylin-Dy(ш) complex by spectrometry using acridine orange as a probe.

    Science.gov (United States)

    Xiong, Xiaoli; Huang, Jianhang; Wang, Xingming

    2014-01-01

    The interaction of a hematoxylin(HE)-Dy(Ш) complex with herring sperm DNA(hsDNA) was studied using acridine orange(AO) as a probe by UV-vis absorption, circular dichroism(CD), fluorescence spectroscopy and viscosity measurements. From the results of the probe experiment, we found that the HE-Dy(Ш) complex could compete with AO for intercalating into hsDNA. The binding constants of the HE-Dy(Ш) complex to hsDNA was obtained by the double reciprocal method and indicated that the affinity between hsDNA and the complex is weaker than that between hsDNA and classical intercalators. The thermodynamic parameters(ΔH°, ΔG°, ΔS°) were calculated from the UV-vis absorption data measured at two different temperatures. Further experimental results suggested that there exist groove binding and partial intercalation binding between hsDNA and HE-Dy(Ш) complex.

  4. Rapid detection of chromosome 18 copy number in buccal smears using DNA probes and FISH

    Energy Technology Data Exchange (ETDEWEB)

    Harris, C.; Nunez, M. [Univ. of Wisconsin, WI (United States); Giraldez, R. [ONCOR, Inc., Gaithersburg, MD (United States)

    1994-09-01

    Rapid diagnosis of trisomy 18 in newborns is often critical to clinical management decisions that must be made in a minimum of time. DNA probes combined with FISH can be used to accurately to determine the copy number of chromosome 18 in interphase cells. We have used the D18Z1 alpha satellite DNA probe to determine signal frequency in normal, previously karyotyped subjects, 12 females and 6 males. We also present one clinical case of trisomy 18, confirmed by karyotype, for comparison to the results obtained from normal subjects. Buccal smears, unlike cytogenetic preparations from peripheral blood, are quite resistant to penetration of probes and detection reagents resulting in higher levels of false monosomy. We have studied 19 individuals and have obtained consistent FISH results, ranging from 64 to 90% disomy. False monosomy rates ranged from 10 to 36%, while false trisomy or tetrasomy was less than 1% in all samples. High rates of false monosomy make this test questionable for detection of low order mosaicism for monosomy, but the extremely low false hyperploidy rate suggests that this is a dependable procedure for detection of trisomy 18, enabling the use of buccal epithelium which can be collected easily from even premature and tiny infants.

  5. Detection of Hepatitis B Virus M204I Mutation by Quantum Dot-Labeled DNA Probe

    Directory of Open Access Journals (Sweden)

    Cheng Zhang

    2017-04-01

    Full Text Available Quantum dots (QDs are semiconductor nanoparticles with a diameter of less than 10 nm, which have been widely used as fluorescent probes in biochemical analysis and vivo imaging because of their excellent optical properties. Sensitive and convenient detection of hepatitis B virus (HBV gene mutations is important in clinical diagnosis. Therefore, we developed a sensitive, low-cost and convenient QDs-mediated fluorescent method for the detection of HBV gene mutations in real serum samples from chronic hepatitis B (CHB patients who had received lamivudine or telbivudine antiviral therapy. We also evaluated the efficiency of this method for the detection of drug-resistant mutations compared with direct sequencing. In CHB, HBV DNA from the serum samples of patients with poor response or virological breakthrough can be hybridized to probes containing the M204I mutation to visualize fluorescence under fluorescence microscopy, where fluorescence intensity is related to the virus load, in our method. At present, the limits of the method used to detect HBV genetic variations by fluorescence quantum dots is 103 IU/mL. These results show that QDs can be used as fluorescent probes to detect viral HBV DNA polymerase gene variation, and is a simple readout system without complex and expensive instruments, which provides an attractive platform for the detection of HBV M204I mutation.

  6. Meat speciation by restriction fragment length polymorphism analysis using an α-actin cDNA probe.

    Science.gov (United States)

    Fairbrother, K S; Hopwood, A J; Lockley, A K; Bardsley, R G

    1998-09-01

    Classical DNA fingerprinting is based on separation of DNA restriction fragments by electrophoresis and hybridisation to nucleic acid probes containing repetitive nucleotide sequences. The use of such mini- or micro-satellite probes tends to yield patterns specific to an individual rather than to a species, hence their value in forensic analysis but general unsuitability for meat speciation. In the present study, a cDNA probe based on conserved sequences contained in members of the actin multigene family has been evaluated for potential application in meat speciation. Genomic DNA was extracted from muscle and digested with BamHI before electrophoresis and hybridisation to a murine α-actin cDNA probe. Beef, pork, lamb, horse, chicken and fish DNA restriction fragments formed characteristic 'fingerprints' which were reproducible and varied sufficiently to allow discrimination even between closely-related species. However no major differences were seen between individuals of the same breed or between different breeds within a species. When DNA obtained from fresh tissue and also from meat heated at 120 °C was analysed, the gel patterns were essentially the same. An attractive feature of this approach is that it employs a single cross-reacting probe and set of conditions, and gives different patterns with all species so far studied. This simplicity suggests applications in meat speciation or related areas of biology.

  7. Ultrafast excited-state dynamics at interfaces: fluorescent DNA probes at the dodecane/water interface

    Science.gov (United States)

    Licari, Giuseppe; Vauthey, Eric

    2015-08-01

    Although the interfaces between two isotropic media are of primary importance in many areas of science and technology, their properties are only partially understood. Our strategy to obtain an insight into these properties is to investigate the ultrafast excited-state dynamics of environment-sensitive molecular probes at liquid interfaces using time-resolved surface second harmonic generation, and to compare it with the dynamics of the same molecules in bulk solutions. Additionally, this approach gives rich information on how the chemical reactivity may change when going from the bulk phase to the interface. This is illustrated by an investigation performed with a series of fluorescent DNA probes at the dodecane/water interface without and with the presence of DNA in the aqueous phase. Substantial differences in the conformation of these cyanine dyes (aggregated or not) and in the excited-state dynamics are observed when going from bulk solutions to the interface. Moreover, the presence of double-stranded DNA in the aqueous phase induces some chirality at the interface.

  8. DNA-based stable isotope probing enables the identification of active bacterial endophytes in potatoes.

    Science.gov (United States)

    Rasche, Frank; Lueders, Tillmann; Schloter, Michael; Schaefer, Sabine; Buegger, Franz; Gattinger, Andreas; Hood-Nowotny, Rebecca C; Sessitsch, Angela

    2009-03-01

    A (13)CO2 (99 atom-%, 350 ppm) incubation experiment was performed to identify active bacterial endophytes in two cultivars of Solanum tuberosum, cultivars Desirée and Merkur. We showed that after the assimilation and photosynthetic transformation of (13)CO2 into (13)C-labeled metabolites by the plant, the most directly active, cultivar specific heterotrophic endophytic bacteria that consume these labeled metabolite scan be identified by DNA stable isotope probing (DNA-SIP).Density-resolved DNA fractions obtained from SIP were subjected to 16S rRNA gene-based community analysis using terminal restriction fragment length polymorphism analysis and sequencing of generated gene libraries.Community profiling revealed community compositions that were dominated by plant chloroplast and mitochondrial 16S rRNA genes for the 'light' fractions of (13)CO2-incubated potato cultivars and of potato cultivars not incubated with (13)CO2. In the 'heavy' fractions of the (13)CO2-incubated endophyte DNA, a bacterial 492-bp terminal restriction fragment became abundant, which could be clearly identified as Acinetobacter and Acidovorax spp. in cultivars Merkur and Desirée,respectively, indicating cultivar-dependent distinctions in (13)C-label flow. These two species represent two common potato endophytes with known plant-beneficial activities.The approach demonstrated the successful detection of active bacterial endophytes in potato. DNA-SIP therefore offers new opportunities for exploring the complex nature of plant-microbe interactions and plant-dependent microbial metabolisms within the endosphere.

  9. DNA-encapsulated silver nanodots as ratiometric luminescent probes for hypochlorite detection

    Science.gov (United States)

    Park, Soonyoung; Choi, Sungmoon; Yu, Junhua

    2014-03-01

    DNA-encapsulated silver nanodots are noteworthy candidates for bio-imaging probes, thanks to their excellent photophysical properties. The spectral shift of silver nanodot emitters from red to blue shows excellent correlations with the concentration of reactive oxygen species, which makes it possible to develop new types of probes for reactive oxygen species (ROS), such as hypochlorous acid (HOCl), given the outstanding stability of the blue in oxidizing environments. HOCl plays a role as a microbicide in immune systems but, on the other hand, is regarded as a disease contributor. Moreover, it is a common ingredient in household cleaners. There are still great demands to detect HOCl fluxes and their physiological pathways. We introduce a new ratiometric luminescence imaging method based on silver nanodots to sensitively detect hypochlorite. The factors that influence the accuracy of the detection are investigated. Its availability has also been demonstrated by detecting the active component in cleaners.

  10. The attachment and characterization of DNA probes on gallium arsenide-based semiconductor surfaces

    Science.gov (United States)

    Yang, Joonhyuk

    2007-12-01

    Immobilization of nucleic acid molecules on solid surfaces is the core of numerous important technologies in the genomics, disease diagnostics and biosensors applications. The architecture and density of immobilized probe molecules depend on the type of the solid surface on which they are anchored. Even though many different types of surfaces have been studied as substrates for deoxyribonucleic acid (DNA) attachment, the development of a new type of substrate, which is reproducible, stable, highly controlled and easily transferred to practical applications, is still needed. Recent studies have shown that As terminated GaAs-based semiconductors can be used as substrates for immobilized DNA layers. In this study, I aim to understand the attachment of nucleic acid onto the surfaces of As-terminated GaAs-based semiconductors and focus on improving the "brush-structure", which is essential for high quality of biochip based on a DNA layer. Attachment of 8-base and 100-base thiolated ssDNA layers on arsenic terminated GaAs(001) was achieved and characterized. The covalent bonds between the thiolated oligonucleotides with As atoms on the GaAs surface were investigated using x-ray photoelectron spectroscopy (XPS), and the surface morphology was obtained using atomic force microscopy (AFM) and field emission scanning electron microscopy (FESEM). In addition, I studied the effect of DNA length and the presence of a good solvent, such as water, on the oligonucleotides on a GaAs surface. I also investigated the effects of the thiol-based spacer and electrolyte concentration to improve the brush-like structure of the DNA layer. Finally, irradiation effects and AlGaAs resonators have been studied for the applications of DNA brush layer on GaAs as biosensor during the change of attachment probe DNA and hybridization to target DNA. For the 8-base thiolated ssDNA case, AFM results showed that the layer thickness was about ˜2.2 nm in dry mode and increased in wet mode. Replacement

  11. A rapid, cost-effective method of assembly and purification of synthetic DNA probes >100 bp.

    Directory of Open Access Journals (Sweden)

    Michael A Jensen

    Full Text Available Here we introduce a rapid, cost-effective method of generating molecular DNA probes in just under 15 minutes without the need for expensive, time-consuming gel-extraction steps. As an example, we enzymatically concatenated six variable strands (50 bp with a common strand sequence (51 bp in a single pool using Fast-Link DNA ligase to produce 101 bp targets (10 min. Unincorporated species were then filtered out by passing the crude reaction through a size-exclusion column (12 could be achieved with further optimization. Moreover, for large-scale assays, we envision this method to be fully automated with the use of robotics such as the Biomek FX; here, potentially thousands of samples could be pooled, ligated and purified in either a 96, 384 or 1536-well platform in just minutes.

  12. Reactive Microcontact Printing of DNA Probes on (DMA-NAS-MAPS) Copolymer-Coated Substrates for Efficient Hybridization Platforms.

    Science.gov (United States)

    Castagna, Rossella; Bertucci, Alessandro; Prasetyanto, Eko Adi; Monticelli, Marco; Conca, Dario Valter; Massetti, Matteo; Sharma, Parikshit Pratim; Damin, Francesco; Chiari, Marcella; De Cola, Luisa; Bertacco, Riccardo

    2016-04-05

    High-performing hybridization platforms fabricated by reactive microcontact printing of DNA probes are presented. Multishaped PDMS molds are used to covalently bind oligonucleotides over a functional copolymer (DMA-NAS-MAPS) surface. Printed structures with minimum width of about 1.5 μm, spaced by 10 μm, are demonstrated, with edge corrugation lower than 300 nm. The quantification of the immobilized surface probes via fluorescence imaging gives a remarkable concentration of 3.3 × 10(3) oligonucleotides/μm(2), almost totally active when used as probes in DNA-DNA hybridization assays. Indeed, fluorescence and atomic force microscopy show a 95% efficiency in target binding and uniform DNA hybridization over printed areas.

  13. Multiplex Ligation-Dependent Probe Amplification Technique for Copy Number Analysis on Small Amounts of DNA Material

    DEFF Research Database (Denmark)

    Sørensen, Karina; Andersen, Paal; Larsen, Lars;

    2008-01-01

    The multiplex ligation-dependent probe amplification (MLPA) technique is a sensitive technique for relative quantification of up to 50 different nucleic acid sequences in a single reaction, and the technique is routinely used for copy number analysis in various syndromes and diseases. The aim...... of the study was to exploit the potential of MLPA when the DNA material is limited. The DNA concentration required in standard MLPA analysis is not attainable from dried blood spot samples (DBSS) often used in neonatal screening programs. A novel design of MLPA probes has been developed to permit for MLPA...... analysis on small amounts of DNA. Six patients with congenital adrenal hyperplasia (CAH) were used in this study. DNA was extracted from both whole blood and DBSS and subjected to MLPA analysis using normal and modified probes. Results were analyzed using GeneMarker and manual Excel analysis. A total...

  14. Research trend of fiber-optic DNA biosensor%光纤DNA生物传感器的研究动向

    Institute of Scientific and Technical Information of China (English)

    黄琛; 黄智伟

    2002-01-01

    DNA生物传感器是分子生物学与微电子学、电化学、光学等相结合的产物,光纤DNA生物传感器是近年DNA 生物传感器中发展最快的一类.介绍了光纤DNA生物传感器的结构原理及研究动向.

  15. Dissecting the effect of anions on Hg2+ detection using a FRET based DNA probe.

    Science.gov (United States)

    Kiy, Mehmet Murat; Zaki, Ahmed; Menhaj, Arsalsan Beg; Samadi, Azadeh; Liu, Juewen

    2012-08-07

    Many biosensors have been developed to detect Hg(2+) using thymine-rich DNA. While sensor response to various cations is often studied to demonstrate selectivity, the effect of anions has been largely overlooked. Anions may compete with DNA for metal binding and thus produce a false negative result. Anions cannot be added alone; the cation part of a salt may cause DNA compaction and other effects, obscuring the role of anions. We find that the sensitivity of a FRET-based Hg(2+) probe is independent of Na(+) concentration. Therefore, by using various sodium salts, any change in sensitivity can be attributed solely to the effect of anions. Halide salts, sulfides, and amines are strong inhibitors; anions containing oxo or hydroxyl groups (e.g. nitrate, sulfate, phosphate, carbonate, acetate, and citrate) do not interfere with Hg(2+) detection even at 100 mM concentration. Mercury hydrolysis and its diffusion into polypropylene containers can also strongly affect the detection results. We conclude that thymine-rich DNA should be useful for Hg(2+) detection in many environmental water samples.

  16. LINE-1 repetitive DNA probes for species-specific cloning from Mus spretus and Mus domesticus genomes.

    Science.gov (United States)

    Rikke, B A; Hardies, S C

    1991-12-01

    Mus domesticus and Mus spretus mice are closely related subspecies. For genetic investigations involving hybrid mice, we have developed a set of species-specific oligonucleotide probes based on the detection of LINE-1 sequence differences. LINE-1 is a repetitive DNA family whose many members are interspersed among the genes. In this study, library screening experiments were used to fully characterize the species specificity of four M. domesticus LINE-1 probes and three M. spretus LINE-1 probes. It was found that the nucleotide differences detected by the probes define large, species-specific subfamilies. We show that collaborative use of such probes can be employed to selectively detect thousands of species-specific library clones. Consequently, these probes could be exploited to monitor and access almost any given species-specific region of interest within hybrid genomes.

  17. Genetic effect of A-bomb radiation- Analysis of minisatellite regions detected by DNA fingerprint probe

    Energy Technology Data Exchange (ETDEWEB)

    Kodaira, Mieko [Radiation Effects Research Foundation, Hiroshima (Japan)

    1999-06-01

    In author's laboratory, screening of mutation in germ cells of A-bomb survivors is under investigation with use of 8 single-locus minisatellite probes and no increase in mutation rate has been detected hitherto. This paper reported results of screening on the minisatellite region, which consisting of short repeated base sequence, using a DNA fingerprint probe for 33.15 core sequence. Subjects were 50 A-bomb survivor families exposed to mean dose of 1.9 Sv (exposed group) or 0 Gy (control), having 64 or 60 children, respectively. DNA was extracted from their B cells established by EB virus and subjected to agarose-gel electrophoresis followed by southern blotting with some improvements for fingerprinting. On the fingerprints, numbers of the band detected in regions of >3.5 kb were 1080 in children of the exposed group (16.9/child) and 1024 (17.1) in the control group, indicating no detectable effect of exposure on the germ cell mutation rate in the region.(K.H.)

  18. Spectroscopic studies on the interaction mechanisms of safranin T with herring sperm DNA using acridine orange as a fluorescence probe.

    Science.gov (United States)

    Long, Jun; Wang, Xing-ming; Xu, Dong-ling; Ding, Li-sheng

    2014-03-01

    Under the condition of physiological pH environment (pH = 7.40), the interactions of safranin T (ST) with herring sperm DNA were studied by means of spectral methods using acridine orange (AO) as a fluorescence probe. The spectroscopic characteristics of DNA-AO in the case of ST (along with the increase of concentration) were observed in an aqueous medium. The binding constants for ST stranded DNA and competitive bindings of ST interacting with DNA-AO systems were examined by fluorescence spectra, and the binding mechanism of ST with DNA was researched via viscosity measurements. All the testimony manifested that bonding modes between ST and DNA were evidenced to be intercalative binding and electrostatic binding, and the combining constant of ST with DNA was obtained. The binding of ST to DNA was driven by entropy and enthalpy through the calculated thermodynamic parameters (Δr Hm (Ө), Δr Sm and Δr Gm (Ө)).

  19. Cationic lipids and cationic ligands induce DNA helix denaturation: detection of single stranded regions by KMnO4 probing.

    Science.gov (United States)

    Prasad, T K; Gopal, Vijaya; Rao, N Madhusudhana

    2003-09-25

    Cationic lipids and cationic polymers are widely used in gene delivery. Using 1,2-dioleoyl-3-trimethylammonium-propane (DOTAP) as a cationic lipid, we have investigated the stability of the DNA in DOTAP:DNA complexes by probing with potassium permanganate (KMnO4). Interestingly, thymidines followed by a purine showed higher susceptibility to cationic ligand-mediated melting. Similar studies performed with other water-soluble cationic ligands such as polylysine, protamine sulfate and polyethyleneimine also demonstrated melting of the DNA but with variations. Small cations such as spermine and spermidine and a cationic detergent, cetyl trimethylammonium bromide, also rendered the DNA susceptible to modification by KMnO4. The data presented here provide direct proof for melting of DNA upon interaction with cationic lipids. Structural changes subsequent to binding of cationic lipids/ligands to DNA may lead to instability and formation of DNA bubbles in double-stranded DNA.

  20. Kelvin probe force microscopy of DNA-capped nanoparticles for single-nucleotide polymorphism detection

    Science.gov (United States)

    Lee, Hyungbeen; Lee, Sang Won; Lee, Gyudo; Lee, Wonseok; Lee, Jeong Hoon; Hwang, Kyo Seon; Yang, Jaemoon; Lee, Sang Woo; Yoon, Dae Sung

    2016-07-01

    Kelvin probe force microscopy (KPFM) is a robust toolkit for profiling the surface potential (SP) of biomolecular interactions between DNAs and/or proteins at the single molecule level. However, it has often suffered from background noise and low throughput due to instrumental or environmental constraints, which is regarded as limiting KPFM applications for detection of minute changes in the molecular structures such as single-nucleotide polymorphism (SNP). Here, we show KPFM imaging of DNA-capped nanoparticles (DCNP) that enables SNP detection of the BRCA1 gene owing to sterically well-adjusted DNA-DNA interactions that take place within the confined spaces of DCNP. The average SP values of DCNP interacting with BRCA1 SNP were found to be lower than the DCNP reacting with normal (non-mutant) BRCA1 gene. We also demonstrate that SP characteristics of DCNP with different substrates (e.g., Au, Si, SiO2, and Fe) provide us with a chance to attenuate or augment the SP signal of DCNP without additional enhancement of instrumentation capabilities.Kelvin probe force microscopy (KPFM) is a robust toolkit for profiling the surface potential (SP) of biomolecular interactions between DNAs and/or proteins at the single molecule level. However, it has often suffered from background noise and low throughput due to instrumental or environmental constraints, which is regarded as limiting KPFM applications for detection of minute changes in the molecular structures such as single-nucleotide polymorphism (SNP). Here, we show KPFM imaging of DNA-capped nanoparticles (DCNP) that enables SNP detection of the BRCA1 gene owing to sterically well-adjusted DNA-DNA interactions that take place within the confined spaces of DCNP. The average SP values of DCNP interacting with BRCA1 SNP were found to be lower than the DCNP reacting with normal (non-mutant) BRCA1 gene. We also demonstrate that SP characteristics of DCNP with different substrates (e.g., Au, Si, SiO2, and Fe) provide us with a

  1. Isolation and characterization of DNA probes from a flow-sorted human chromosome 8 library that detect restriction fragment length polymorphism (RFLP).

    OpenAIRE

    Wood, S.; Starr, T V; Shukin, R J

    1986-01-01

    We have used a recombinant DNA library constructed from flow-sorted human chromosome 8 as a source of single-copy human probes. These probes have been screened for restriction fragment length polymorphism (RFLP) by hybridization to Southern transfers of genomic DNA from five unrelated individuals. We have detected six RFLPs distributed among four probes after screening 741 base pairs for restriction site variation. These RFLPs all behave as codominant Mendelian alleles. Two of the probes dete...

  2. The interaction of taurine-salicylaldehyde Schiff base copper(II) complex with DNA and the determination of DNA using the complex as a fluorescence probe

    Science.gov (United States)

    Zhang, Xiaoyan; Wang, Yong; Zhang, Qianru; Yang, Zhousheng

    2010-09-01

    The interaction of taurine-salicylaldehyde Schiff base copper(II) (Cu(TSSB) 22+) complex with DNA was explored by using UV-vis, fluorescence spectrophotometry, and voltammetry. In pH 7.4 Tris-HCl buffer solution, the binding constant of the Cu(TSSB) 22+ complex interaction with DNA was 3.49 × 10 4 L mol -1. Moreover, due to the fluorescence enhancing of Cu(TSSB) 22+ complex in the presence of DNA, a method for determination of DNA with Cu(TSSB) 22+ complex as a fluorescence probe was developed. The fluorescence spectra indicated that the maximum excitation and emission wavelength were 389 nm and 512 nm, respectively. Under optimal conditions, the calibration graphs are linear over the range of 0.03-9.03 μg mL -1 for calf thymus DNA (CT-DNA), 0.10-36 μg mL -1 for yeast DNA and 0.01-10.01 μg mL -1 for salmon DNA (SM-DNA), respectively. The corresponding detection limits are 7 ng mL -1 for CT-DNA, 3 ng mL -1 for yeast DNA and 3 ng mL -1 for SM-DNA. Using this method, DNA in synthetic samples was determined with satisfactory results.

  3. Direct fluorescence in situ hybridization on human metaphase chromosomes using quantum dot-platinum labeled DNA probes

    Energy Technology Data Exchange (ETDEWEB)

    Hwang, Gyoyeon [Chemical Kinomics Research Center, Future Convergence Research Division, Korea Institute of Science and Technology, Hwarangno 14-gil 5, Seongbuk-gu, Seoul 136-791 (Korea, Republic of); Biological Chemistry, Korea University of Science and Technology, 217, Gajeong-ro, Yuseong-gu, Deajeon (Korea, Republic of); Lee, Hansol [Chemical Kinomics Research Center, Future Convergence Research Division, Korea Institute of Science and Technology, Hwarangno 14-gil 5, Seongbuk-gu, Seoul 136-791 (Korea, Republic of); Lee, Jiyeon, E-mail: jylee@kist.re.kr [Chemical Kinomics Research Center, Future Convergence Research Division, Korea Institute of Science and Technology, Hwarangno 14-gil 5, Seongbuk-gu, Seoul 136-791 (Korea, Republic of); Biological Chemistry, Korea University of Science and Technology, 217, Gajeong-ro, Yuseong-gu, Deajeon (Korea, Republic of)

    2015-11-13

    The telomere shortening in chromosomes implies the senescence, apoptosis, or oncogenic transformation of cells. Since detecting telomeres in aging and diseases like cancer, is important, the direct detection of telomeres has been a very useful biomarker. We propose a telomere detection method using a newly synthesized quantum dot (QD) based probe with oligonucleotide conjugation and direct fluorescence in situ hybridization (FISH). QD-oligonucleotides were prepared with metal coordination bonding based on platinum-guanine binding reported in our previous work. The QD-oligonucleotide conjugation method has an advantage where any sequence containing guanine at the end can be easily bound to the starting QD-Pt conjugate. A synthesized telomeric oligonucleotide was bound to the QD-Pt conjugate successfully and this probe hybridized specifically on the telomere of fabricated MV-4-11 and MOLT-4 chromosomes. Additionally, the QD-telomeric oligonucleotide probe successfully detected the telomeres on the CGH metaphase slide. Due to the excellent photostability and high quantum yield of QDs, the QD-oligonucleotide probe has high fluorescence intensity when compared to the organic dye-oligonucleotide probe. Our QD-oligonucleotide probe, conjugation method of this QD probe, and hybridization protocol with the chromosomes can be a useful tool for chromosome painting and FISH. - Highlights: • We prepared a probe linked between QD and telomeric oligonucleotide with platinum-guanine bonding. • Telomeres were detected by our new telomere probes successfully in three different human metaphase chromosomes. • QDPt-DNA probe has high fluorescence intensity in comparison with organic dye-DNA probe.

  4. Novel Phenanthrene-Degrading Bacteria Identified by DNA-Stable Isotope Probing.

    Directory of Open Access Journals (Sweden)

    Longfei Jiang

    Full Text Available Microorganisms responsible for the degradation of phenanthrene in a clean forest soil sample were identified by DNA-based stable isotope probing (SIP. The soil was artificially amended with either 12C- or 13C-labeled phenanthrene, and soil DNA was extracted on days 3, 6 and 9. Terminal restriction fragment length polymorphism (TRFLP results revealed that the fragments of 219- and 241-bp in HaeIII digests were distributed throughout the gradient profile at three different sampling time points, and both fragments were more dominant in the heavy fractions of the samples exposed to the 13C-labeled contaminant. 16S rRNA sequencing of the 13C-enriched fraction suggested that Acidobacterium spp. within the class Acidobacteria, and Collimonas spp. within the class Betaproteobacteria, were directly involved in the uptake and degradation of phenanthrene at different times. To our knowledge, this is the first report that the genus Collimonas has the ability to degrade PAHs. Two PAH-RHDα genes were identified in 13C-labeled DNA. However, isolation of pure cultures indicated that strains of Staphylococcus sp. PHE-3, Pseudomonas sp. PHE-1, and Pseudomonas sp. PHE-2 in the soil had high phenanthrene-degrading ability. This emphasizes the role of a culture-independent method in the functional understanding of microbial communities in situ.

  5. Identification of the autotrophic denitrifying community in nitrate removal reactors by DNA-stable isotope probing.

    Science.gov (United States)

    Xing, Wei; Li, Jinlong; Cong, Yuan; Gao, Wei; Jia, Zhongjun; Li, Desheng

    2017-04-01

    Autotrophic denitrification has attracted increasing attention for wastewater with insufficient organic carbon sources. Nevertheless, in situ identification of autotrophic denitrifying communities in reactors remains challenging. Here, a process combining micro-electrolysis and autotrophic denitrification with high nitrate removal efficiency was presented. Two batch reactors were fed organic-free nitrate influent, with H(13)CO3(-) and H(12)CO3(-) as inorganic carbon sources. DNA-based stable-isotope probing (DNA-SIP) was used to obtain molecular evidence for autotrophic denitrifying communities. The results showed that the nirS gene was strongly labeled by H(13)CO3(-), demonstrating that the inorganic carbon source was assimilated by autotrophic denitrifiers. High-throughput sequencing and clone library analysis identified Thiobacillus-like bacteria as the most dominant autotrophic denitrifiers. However, 88% of nirS genes cloned from the (13)C-labeled "heavy" DNA fraction showed low similarity with all culturable denitrifiers. These findings provided functional and taxonomical identification of autotrophic denitrifying communities, facilitating application of autotrophic denitrification process for wastewater treatment.

  6. A competition assay for DNA binding using the fluorescent probe ANS.

    Science.gov (United States)

    Taylor, Ian A; Kneale, G Geoff

    2009-01-01

    Fluorescence spectroscopy is a technique frequently employed to study protein-nucleic acid interactions. Often, the intrinsic fluorescence emission spectrum of tryptophan residues in a nucleic-acid-binding protein is strongly perturbed upon interaction with a target DNA or RNA. These spectral changes can then be exploited in order to construct binding isotherms and the extract equilibrium association constant together with the stoichiometry of an interaction. However, when a protein contains many tryptophan residues that are not located in the proximity of the nucleic-acid-binding site, changes in the fluorescence emission spectrum may not be apparent or the magnitude too small to be useful. Here, we make use of an extrinsic fluorescence probe, the environmentally sensitive fluorophore 1-anilinonaphthalene-8-sulphonic acid (1,8-ANS). Displacement by DNA of 1,8-ANS molecules from the nucleic-acid-binding site of the Type I modification methylase EcoR124I results in red shifting and an intensity decrease of the 1,8-ANS fluorescence emission spectrum. These spectral changes have been used to investigate the interaction of EcoR124I with DNA target recognition sequences.

  7. THE CLONING OF HRNT-1 USING A COMBINATION OF cDNA LIBRARY SCREENING WITH BIOTIN-LABELED PROBE AND RAPID AMPLIFICATION OF cDNA ENDS

    Institute of Scientific and Technical Information of China (English)

    2001-01-01

    Objective: To clone the human counterpart of rat ZA73, EST cloned from rat tracheal epithelial (RTE) neoplastic transformed cell model induced by (a-particles radiation by using mRNA differential display. Methods: According to the sequence of rat ZA73, a probe was biotin-labeled to screen human cDNA library, and then the gene sequence was extended by RACE (rapid amplification of cDNA ends). Result: Human gene HRNT-1 (GenBank Accession Number: AF223393) is 4.256 kb in length, with an ORF located in the region between 254 and 3013 bp. 5' UTS (untranslated sequences) is 253 bp, 3' UTS is 1243 bp. Conclusion: The combination of cDNA library screening with biotin-labeled probes and RACE is an effective method to clone full-length cDNA, especially for sequences longer than 2 kb.

  8. Effect of a Dual Charge on the DNA-Conjugated Redox Probe on DNA Sensing by Short Hairpin Beacons Tethered to Gold Electrodes.

    Science.gov (United States)

    Kékedy-Nagy, László; Shipovskov, Stepan; Ferapontova, Elena E

    2016-08-16

    Charges of redox species can critically affect both the interfacial state of DNA and electrochemistry of DNA-conjugated redox labels and, as a result, the electroanalytical performance of those systems. Here, we show that the kinetics of electron transfer (ET) between the gold electrode and methylene blue (MB) label conjugated to a double-stranded (ds) DNA tethered to gold strongly depend on the charge of the MB molecule, and that affects the performance of genosensors exploiting MB-labeled hairpin DNA beacons. Positively charged MB binds to dsDNA via electrostatic and intercalative/groove binding, and this binding allows the DNA-mediated electrochemistry of MB intercalated into the duplex and, as a result, a complex mode of the electrochemical signal change upon hairpin hybridization to the target DNA, dominated by the "on-off" signal change mode at nanomolar levels of the analyzed DNA. When MB bears an additional carboxylic group, the negative charge provided by this group prevents intimate interactions between MB and DNA, and then the ET in duplexes is limited by the diffusion of the MB-conjugated dsDNA (the phenomenon first shown in Farjami , E. ; Clima , L. ; Gothelf , K. ; Ferapontova , E. E. Anal. Chem. 2011 , 83 , 1594 ) providing the robust "off-on" nanomolar DNA sensing. Those results can be extended to other intercalating redox probes and are of strategic importance for design and development of electrochemical hybridization sensors exploiting DNA nanoswitchable architectures.

  9. Detection of beer spoilage bacteria Pectinatus and Megasphaera with acridinium ester labelled DNA probes using a hybridisation protection assay.

    Science.gov (United States)

    Paradh, A D; Hill, A E; Mitchell, W J

    2014-01-01

    DNA probes specific for rRNA of selected target species were utilised for the detection of beer spoilage bacteria of the genera Pectinatus and Megasphaera using a hybridisation protection assay (HPA). All the probes were modified during synthesis by addition of an amino linker arm at the 5' end or were internally modified by inserting an amine modified thymidine base. Synthesised probes then were labelled with acridinium ester (AE) and purified using reverse phase HPLC. The internally AE labelled probes were able to detect target RNA within the range of 0.016-0.0032pmol. All the designed probes showed high specificity towards target RNA and could detect bacterial contamination within the range of ca. 5×10(2)1×10(3) CFU using the HPA. The developed assay was also compatible with MRS, NBB and SMMP beer enrichment media, routinely used in brewing laboratories.

  10. Potential use of buccal smears for rapid diagnosis of autosomal trisomy or chromosomal sex in newborn infants using DNA probes

    Energy Technology Data Exchange (ETDEWEB)

    Harris, C.; Clark, K.; Lazarski, K. [Univ. of Wisconsin, Madison, WI (United States); Wilkerson, C. [Univ. of Wisconsin Medical School, Madison, WI (United States); Meisner, L. [Univ. of Wisconsin, Madison, WI (United States)]|[Univ. of Wisconsin Medical School, Madison, WI (United States)

    1994-12-01

    Buccal smears from 3 women and 1 man were probed with alpha satellite DNA probes for chromosomes 8, 18, X, and Y. Buccal smears were also collected from an adolescent phenotypic female with uterine agenesis, as well as from newborn infants with suspected trisomy 18 and trisomy 21. The clinical cases were confirmed with conventional cytogenetic studies of peripheral lymphocytes. Overall probe efficiency at detecting expected chromosome number in interphase cells was found to be 71% {+-} 6.8%. Higher than expected n-1 signal numbers may be due to karyopyknotic intermediate epithelial cells present in all collected samples. Overall probe efficiency was found to be consistent using alpha satellite and cosmid probes, both of which accurately reflected the modal copy number of the target chromosomes. False trisomy was less than 1%. This study suggests DNA probes can be used in buccal smears for rapid diagnosis of trisomies and chromosomal sex in newborns, but because of high rates of false hydropoploid signals, probed buccal smear specimens may not be accurate at diagnosing mosaicism. 9 refs., 2 figs., 1 tab.

  11. Cyanotryptophans as Novel Fluorescent Probes for Studying Protein Conformational Changes and DNA-Protein Interaction.

    Science.gov (United States)

    Talukder, Poulami; Chen, Shengxi; Roy, Basab; Yakovchuk, Petro; Spiering, Michelle M; Alam, Mohammad P; Madathil, Manikandadas M; Bhattacharya, Chandrabali; Benkovic, Stephen J; Hecht, Sidney M

    2015-12-29

    Described herein are the syntheses and photophysical characterization of three novel cyanotryptophans, and their efficient incorporation into proteins as fluorescent probes. Photophysical characteristics indicated that each was significantly brighter and red-shifted in fluorescence emission relative to tryptophan. Each analogue was used to activate a suppressor tRNA transcript and was incorporated with good efficiency into two different positions (Trp22 and Trp74) of Escherichia coli dihydrofolate reductase (ecDHFR). The Trp analogues could be monitored selectively in the presence of multiple native Trp residues in DHFR. 6-CNTrp (A) formed an efficient Förster resonance energy transfer (FRET) pair with l-(7-hydroxycoumarin-4-yl)ethylglycine (HCO, D) at position 17. Further, 6-CNTrp (A) was incorporated into two DNA binding proteins, including the Klenow fragment of DNA polymerase I and an RNA recognition motif (RRM2) of heterogeneous nuclear ribonucleoprotein L-like (hnRNP LL). Using these proteins, we demonstrated the use of FRET involving A as a fluorescence donor and benzo[g]quinazoline-2,4-(1H,3H)-dione 2'-deoxyriboside (Tf) or 4-aminobenzo[g]quinazoline-2-one 2'-deoxyriboside (Cf) as fluorescent acceptors to study the binding interaction of the Klenow fragment with duplex DNA oligomers (labeled with Tf), or the domain-specific association between hnRNP LL and the BCL2 i-motif DNA (labeled with Cf). Thus, the non-natural amino acid could be used as a FRET partner for studying protein-nucleic acid interactions. Together, these findings demonstrate the potential utility of 6-CNTrp (A) as a fluorescence donor for the study of protein conformational events.

  12. Nutrient amendments in soil DNA stable isotope probing experiments reduce the observed methanotroph diversity.

    Science.gov (United States)

    Cébron, Aurélie; Bodrossy, Levente; Stralis-Pavese, Nancy; Singer, Andrew C; Thompson, Ian P; Prosser, James I; Murrell, J Colin

    2007-02-01

    Stable isotope probing (SIP) can be used to analyze the active bacterial populations involved in a process by incorporating 13C-labeled substrate into cellular components such as DNA. Relatively long incubation times are often used with laboratory microcosms in order to incorporate sufficient 13C into the DNA of the target organisms. Addition of nutrients can be used to accelerate the processes. However, unnatural concentrations of nutrients may artificially change bacterial diversity and activity. In this study, methanotroph activity and diversity in soil was examined during the consumption of 13CH4 with three DNA-SIP experiments, using microcosms with natural field soil water conditions, the addition of water, and the addition of mineral salts solution. Methanotroph population diversity was studied by targeting 16S rRNA and pmoA genes. Clone library analyses, denaturing gradient gel electrophoresis fingerprinting, and pmoA microarray hybridization analyses were carried out. Most methanotroph diversity (type I and type II methanotrophs) was observed in non-amended SIP microcosms. Although this treatment probably best reflected the in situ environmental conditions, one major disadvantage of this incubation was that the incorporation of 13CH4 was slow and some cross-feeding of 13C occurred, thereby leading to labeling of nonmethanotroph microorganisms. Conversely, microcosms supplemented with mineral salts medium exhibited rapid consumption of 13CH4, resulting in the labeling of a less diverse population of only type I methanotrophs. DNA-SIP incubations using water-amended microcosms yielded faster incorporation of 13C into active methanotrophs while avoiding the cross-feeding of 13C.

  13. The Preparation of Magnetic Silica Nanospheres and Incorporation of CdSe/ZnS Quantum Dots-DNA Probe.

    Science.gov (United States)

    Do, Youngjin; Kim, Jongsung

    2016-03-01

    Silica nanospheres containing magnetic particles were prepared, and CdSe/ZnS QDs functionalized with carboxyl group were incorporated into the silica nanospheres by EDC/NHS coupling reaction. The silica nanospheres were prepared by a co-precipitation of ferrous and ferric solutions followed by the sol-gel reaction of TEOS (tetraethoxysilane) and APTES (3-aminopropyltriethoxysilane) using base catalyst. The size of magnetic silica nanospheres was confirmed by Transmission electron microscope (TEM). Thiol group modified single stranded oligonucleotides were immobilized on the surface of QDs and fluorescence quenching by intercalation dye (TOTO-3) after hybridization with target oligonucleotide was observed. The fluorescence from QDs could be quenched by intercalating dye (TOTO-3) after hybridization of target DNA to probe DNA. This shows that the magnetic silica-QD-DNA probe can be used to detect specific DNA.

  14. Mitochondrial DNA variation in the grasshopper Sinipta dalmani: application of long-PCR to the development of a homologous probe.

    Science.gov (United States)

    Pensel, S M; Vilardi, J C; Remis, M I

    2005-12-01

    RFLP analysis of mtDNA in natural populations is a valuable tool for phylogeographic and population genetic studies. The amplification of long DNA fragments using universal primers may contribute to the development of novel homologous probes in species for which no previous genomic information is available. Here we report how we obtained the complete mtDNA genome of Sinipta dalmani (Orthoptera) in 2 fragments (7 and 9 kb) using primers of conserved regions. The specificity of the PCR reactions was ultimately confirmed by several lines of evidence. These fragments were used as a probe for a mtDNA RFLP study in S. dalmani that analyzed the pattern of haplotype distribution and nucleotide diversity within and among chromosomally differentiated natural populations. Our results suggest that the restriction in gene flow detected at the molecular level may explain the chromosome differentiation detected previously and the maintenance of chromosome polymorphism in some areas of S. dalmani geographic distribution.

  15. Evaluation of a colorimetric Babesia bigemina-DNA probe within an epidemiological survey

    Directory of Open Access Journals (Sweden)

    Juan A. Ramos

    1992-01-01

    Full Text Available An epidemiological survey was conducted in south east Mexico, in an effort to establish the serological reactivity and carrier status to Babesia bigemina of an indigenous cattle population. The prevalance was obtained through the Indirect Fluorescent Antibody Test (IFAT, using an in vitro culture-derived B. bigemina antigen. A specific, digoxigenin-coupled, ~6kb B. bigemina-DNA probe (BBDP, was used to indicate the presence of the parasite. Serum samples from 925 animals of all ages, were obtained within the three regions (I, II, III of the state of Yucatan and tested by IFAT. In addition, whole blood samples draw from 136 of the same animals of region II were analyzed using the BBDP. Positive IFAT (IFAT+ reactions were observed in 531 sera for a 57% overall prevalence. Regional values were: I = 157 + (56%, II = 266 + (68% and III = 108 + (42%. Only 32 (23% of the blood samples tested with BBDP showed distinctive hybridization signal, in contrast with 100 (73% IFAT + animals. The responses distribution for IFAT vs. BBDP was: +/+ 23, +/- 77, -/+ 9 and -/- 27 respectively. It was found that the analytical sinsitivity of BBDP appears to be low for its utilization is widespread epidemiological surveys. It was considered, however, that the colorimetric probe mifht to be useful to safely detect transmission prone carriers, since it is able to detect parasitemias as low as 0.001%.

  16. Detection of hepatitis B virus DNA by real-time PCR using TaqMan-MGB probe technology

    Institute of Scientific and Technical Information of China (English)

    Jin-Rong Zhao; Yu-Jie Bai; Qing-Hua Zhang; Yan Wan; Ding Li; Xiao-Jun Yan

    2005-01-01

    AIM: To develop a real-time PCR for detecting hepatitis B virus-(HBV) DNA based on TaqMan technology using a new MGB probe.METHODS: Plasmid containing the sequence of X gene (1414-1744 nt) was constructed as HBV-DNA standard for quantitative analysis. A TaqMan-MGB probe between primers for amplification was designed to detect PCR products. The interested sequence contained in the plasmid and in clinical specimens was quantitatively measured.RESULTS: The detection limit of the assay for HBV DNA was 1 genome equivalent per reaction. A linear standard curve was obtained between 100 and 109 DNA copies/reaction (r>0.990). None of the negative control samples showed false-positive reactions in duplicate. HBV DNA was detected in 100% (50/50) of HBV patients with HbeAg, and in 72.0% (36/50) with HBsAg, HBeAb and HBcAb. The coefficient of variation for both intra- and inter-experimental variability demonstrated high reproducibility and accuracy.CONCLUSION: Real-time PCR based on TaqMan-MGB probe technology is an excellent method for detection of HBV DNA.

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

    Directory of Open Access Journals (Sweden)

    Stougaard Magnus

    2007-11-01

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

  18. Intravital imaging of fluorescent markers and FRET probes by DNA tattooing

    Directory of Open Access Journals (Sweden)

    Spencer David M

    2007-01-01

    Full Text Available Abstract Background Advances in fluorescence microscopy and mouse transgenesis have made it possible to image molecular events in living animals. However, the generation of transgenic mice is a lengthy process and intravital imaging requires specialized knowledge and equipment. Here, we report a rapid and undemanding intravital imaging method using generally available equipment. Results By DNA tattooing we transfect keratinocytes of living mice with DNA encoding fluorescent biosensors. Subsequently, the behavior of individual cells expressing these biosensors can be visualized within hours and using conventional microscopy equipment. Using this "instant transgenic" model in combination with a corrected coordinate system, we followed the in vivo behavior of individual cells expressing either FRET- or location-based biosensors for several days. The utility of this approach was demonstrated by assessment of in vivo caspase-3 activation upon induction of apoptosis. Conclusion This "instant skin transgenic" model can be used to follow the in vivo behavior of individual cells expressing either FRET- or location-based probes for several days after tattooing and provides a rapid and inexpensive method for intravital imaging in murine skin.

  19. Probing heterobivalent binding to the endocytic AP-2 adaptor complex by DNA-based spatial screening.

    Science.gov (United States)

    Diezmann, F; von Kleist, L; Haucke, V; Seitz, O

    2015-08-01

    The double helical DNA scaffold offers a unique set of properties, which are particularly useful for studies of multivalency in biomolecular interactions: (i) multivalent ligand displays can be formed upon nucleic acid hybridization in a self-assembly process, which facilitates spatial screening (ii) valency and spatial arrangement of the ligand display can be precisely controlled and (iii) the flexibility of the ligand display can be adjusted by integrating nick sites and unpaired template regions. Herein we describe the use of DNA-based spatial screening for the characterization of the adaptor complex 2 (AP-2), a central interaction hub within the endocytic protein network in clathrin-mediated endocytosis. AP-2 is comprised of a core domain and two, so-called appendage domains, the α- and the β2-ear, which associate with cytoplasmatic proteins required for the formation or maturation of clathrin/AP-2 coated pits. Each appendage domain has two binding grooves which recognize distinct peptide motives with micromolar affinity. This provides opportunities for enhanced interactions with protein molecules that contain two (or more) different peptide motives. To determine whether a particular, spatial arrangement of binding motifs is required for high affinity binding we probed the distance-affinity relationships by means of DNA-programmed spatial screening with self-assembled peptide-DNA complexes. By using trimolecular and tetramolecular assemblies two different peptides were positioned in 2-22 nucleotide distance. The binding data obtained with both recombinant protein in well-defined buffer systems and native AP-2 in brain extract suggests that the two binding sites of the AP-2 α-appendage can cooperate to provide up to 40-fold enhancement of affinity compared to the monovalent interaction. The distance between the two recognized peptide motives was less important provided that the DNA duplex segments were connected by flexible, single strand segments. By

  20. GENETIC DIVERSITY OF TYPHA LATIFOLIA (TYPHACEAE) AND THE IMPACT OF POLLUTANTS EXAMINED WITH TANDEM-REPETITIVE DNA PROBES

    Science.gov (United States)

    Genetic diversity at variable-number-tandem-repeat (VNTR) loci was examined in the common cattail, Typha latifolia (Typhaceae), using three synthetic DNA probes composed of tandemly repeated "core" sequences (GACA, GATA, and GCAC). The principal objectives of this investigation w...

  1. Specific detection of neuronal cell bodies: in situ hybridization with a biotin-labelled neurofilament cDNA probe.

    NARCIS (Netherlands)

    P. Liesi; J-P. Julien (Jean-Pierre); P. Vilja; F.G. Grosveld (Frank); L. Rechardt

    1986-01-01

    textabstractWe have used a biotinylated, 300-nucleotide cDNA probe which encodes the 68,000 MW neurofilament protein to detect neurofilament-specific mRNA in situ. The neurofilament message specifically demonstrates the neuronal cell bodies, in contrast to the usual antibody staining which detects t

  2. Multiplexed microRNA detection using lanthanide-labeled DNA probes and laser ablation inductively coupled plasma mass spectrometry

    DEFF Research Database (Denmark)

    de Bang, Thomas Christian; Shah, Pratik; Cho, Seok Keun

    2014-01-01

    . The narrow size range of miRNAs (20-24 nucleotides) combined with the chemical properties of conventional reporter tags has hampered the development of multiplexed miRNA assays. In this study, we have used lanthanide-labeled DNA probes for the detection of miRNAs on membranes using laser ablation inductively...

  3. A liquid chromatography - mass spectrometry method to measure ¹³C-isotope enrichment for DNA stable-isotope probing.

    Science.gov (United States)

    Auclair, Julie; Lépine, François; Villemur, Richard

    2012-03-01

    DNA stable-isotope probing (DNA-SIP) is a cultivation-independent technique that makes it possible to associate metabolic function and taxonomic identity in a wide range of terrestrial and aquatic environments. In DNA-SIP, DNA is labeled via the assimilation of a labeled growth substrate that is subsequently used to identify microorganisms involved in assimilation of the substrate. However, the labeling time has to be sufficient to obtain labeled DNA but not so long such that cross-feeding of ¹³C-labeled metabolites from the primary consumers to nontarget species can occur. Confirmation that the DNA is isotopically labeled in DNA-SIP assays can be achieved using an isotope ratio mass spectrometer. In this study, we describe the development of a method using liquid chromatography (HPLC) coupled to a quadrupole mass spectrometer (QMS) to measure the ¹³C enrichment of thymine incorporated into DNA in Escherichia coli cultures fed with [¹³C]acetate. The method involved the hydrolysis of DNA extracted from the cultures that released the nucleotides, followed by the separation of the thymine by HPLC on a reverse-phase C₈ column in isocratic elution mode and the detection and quantification of ¹³C-labeled thymine by QMS. To mimic a DNA-SIP assay, a DNA mixture was made using ¹³C-labeled E. coli DNA with DNA extracted from five bacterial species. The HPLC-MS method was able to measure the correct proportion of ¹³C-DNA in the mix. This method can then be used as an alternative to the use of isotope ratio mass spectrometry in DNA-SIP assays.

  4. Ultrafast Hydration Dynamics Probed by Tryptophan at Protein Surface and Protein-DNA Interface

    Science.gov (United States)

    Qin, Yangzhong

    As we all live in a special water planet Earth, the significance of water to life has been universally recognized. The reason why water is so important to life has intrigued many researchers. This dissertation will focus on the ultrafast dynamics of protein surface water and protein-DNA interfacial water which have direct importance to the protein structure and function. Using tryptophan as an intrinsic fluorescence probe, combined with site-directed mutagenesis and ultrafast fluorescence up-conversion spectroscopy, we can achieve single residue spatial resolution and femtosecond temporal resolution. We can also precisely determine the local hydration water dynamics by monitoring the Stokes shift of tryptophan one at a time. Previously, the protein surface hydration has been extensively studied by our group. In this thesis, we will provide more details on the methods we are using to extract the hydration dynamics, and also validate our methods from both experimental and theoretical perspectives. To further interrogate the interfacial water hydration dynamics relative to the protein surface hydration, we studied two DNA polymerases: DNA Polymerase IV (Dpo4) and DNA Polymerase Beta (Pol beta). Both proteins show typical surface hydration pattern with three distinct time components including: (i) the ultrafast sub-picosecond component reflects the bulk type water motion; (ii) a few picoseconds component shows the inner water relaxation mainly corresponding to the local libration and reorientation; (iii) the tens to hundred picoseconds component represents the water-protein coupled motion involving the whole water network reorganization. Dpo4, a loosely DNA binding protein, exhibits very flexible interfacial water which resembles its surface water yet with a significantly reduced ultrafast component. Such dynamic interfacial water not only maintains interfacial flexibility, but also contributes to the low fidelity of the protein. In contrast to the Dpo4, pol beta

  5. Spectroscopic quantification of 5-hydroxymethylcytosine in genomic DNA using boric acid-functionalized nano-microsphere fluorescent probes.

    Science.gov (United States)

    Chen, Hua-Yan; Wei, Jing-Ru; Pan, Jiong-Xiu; Zhang, Wei; Dang, Fu-Quan; Zhang, Zhi-Qi; Zhang, Jing

    2017-05-15

    5-hydroxymethylcytosine (5hmC) is the sixth base of DNA. It is involved in active DNA demethylation and can be a marker of diseases such as cancer. In this study, we developed a simple and sensitive 2-(4-boronophenyl)quinoline-4-carboxylic acid modified poly (glycidyl methacrylate (PBAQA-PGMA) fluorescent probe to detect the 5hmC content of genomic DNA based on T4 β-glucosyltransferase-catalyzed glucosylation of 5hmC. The fluorescence-enhanced intensity recorded from the DNA sample was proportional to its 5-hydroxymethylcytosine content and could be quantified by fluorescence spectrophotometry. The developed probe showed good detection sensitivity and selectivity and a good linear relationship between the fluorescence intensity and the concentration of 5 hmC within a 0-100nM range. Compared with other fluorescence detection methods, this method not only could determine trace amounts of 5 hmC from genomic DNA but also could eliminate the interference of fluorescent dyes and the need for purification. It also could avoid multiple labeling. Because the PBAQA-PGMA probe could enrich the content of glycosyl-5-hydroxymethyl-2-deoxycytidine from a complex ground substance, it will broaden the linear detection range and improve sensitivity. The limit of detection was calculated to be 0.167nM after enrichment. Furthermore, the method was successfully used to detect 5-hydroxymethylcytosine from mouse tissues. Copyright © 2016 Elsevier B.V. All rights reserved.

  6. Experimental mapping of DNA duplex shape enabled by global lineshape analyses of a nucleotide-independent nitroxide probe.

    Science.gov (United States)

    Ding, Yuan; Zhang, Xiaojun; Tham, Kenneth W; Qin, Peter Z

    2014-10-01

    Sequence-dependent variation in structure and dynamics of a DNA duplex, collectively referred to as 'DNA shape', critically impacts interactions between DNA and proteins. Here, a method based on the technique of site-directed spin labeling was developed to experimentally map shapes of two DNA duplexes that contain response elements of the p53 tumor suppressor. An R5a nitroxide spin label, which was covalently attached at a specific phosphate group, was scanned consecutively through the DNA duplex. X-band continuous-wave electron paramagnetic resonance spectroscopy was used to monitor rotational motions of R5a, which report on DNA structure and dynamics at the labeling site. An approach based on Pearson's coefficient analysis was developed to collectively examine the degree of similarity among the ensemble of R5a spectra. The resulting Pearson's coefficients were used to generate maps representing variation of R5a mobility along the DNA duplex. The R5a mobility maps were found to correlate with maps of certain DNA helical parameters, and were capable of revealing similarity and deviation in the shape of the two closely related DNA duplexes. Collectively, the R5a probe and the Pearson's coefficient-based lineshape analysis scheme yielded a generalizable method for examining sequence-dependent DNA shapes.

  7. Highly Sensitive Temperature Sensors Based on Fiber-Optic PWM and Capacitance Variation Using Thermochromic Sensing Membrane.

    Science.gov (United States)

    Khan, Md Rajibur Rahaman; Kang, Shin-Won

    2016-07-09

    In this paper, we propose a temperature/thermal sensor that contains a Rhodamine-B sensing membrane. We applied two different sensing methods, namely, fiber-optic pulse width modulation (PWM) and an interdigitated capacitor (IDC)-based temperature sensor to measure the temperature from 5 °C to 100 °C. To the best of our knowledge, the fiber-optic PWM-based temperature sensor is reported for the first time in this study. The proposed fiber-optic PWM temperature sensor has good sensing ability; its sensitivity is ~3.733 mV/°C. The designed temperature-sensing system offers stable sensing responses over a wide dynamic range, good reproducibility properties with a relative standard deviation (RSD) of ~0.021, and the capacity for a linear sensing response with a correlation coefficient of R² ≈ 0.992 over a wide sensing range. In our study, we also developed an IDC temperature sensor that is based on the capacitance variation principle as the IDC sensing element is heated. We compared the performance of the proposed temperature-sensing systems with different fiber-optic temperature sensors (which are based on the fiber-optic wavelength shift method, the long grating fiber-optic Sagnac loop, and probe type fiber-optics) in terms of sensitivity, dynamic range, and linearity. We observed that the proposed sensing systems have better sensing performance than the above-mentioned sensing system.

  8. Surface modification of poly(dimethylsiloxane) (PDMS) microchannels with DNA capture-probes for potential use in microfluidic DNA analysis systems

    Science.gov (United States)

    Khodakov, Dmitriy A.; Thredgold, Leigh D.; Lenehan, Claire E.; Andersson, Gunther A.; Kobus, Hilton; Ellis, Amanda V.

    2011-12-01

    Poly(dimethylsiloxane) (PDMS) is an elastomeric material used for microfluidic devices and is especially suited to medical and forensic applications. This is due to its relatively low cost, ease of fabrication, excellent optical transmission characteristics and its ability to support electroosmotic flow, required during electrophoretic separations. These aspects combined with its large range of surface modification chemistries, make PDMS an attractive substrate in microfluidic devices for, in particular, DNA separation. Here, we report the successful wet chemical surface modification of PDMS microchannels using a simple three step method to produce an isothiocyanate-terminated surface. Initially, PDMS was oxygen plasma treated to produce a silanol-terminated surface, this was then reacted with 3-aminopropyltriethoxysilane with subsequent reaction of the now amine-terminated surface with p-phenylenediisothiocyanate. Water contact angle measurements both before and after modification showed a reduction in hydrophobicity from 101o for native PDMS to 94o for the isothiocyante-terminated PDMS. The isothiocyanate-terminated surface was then coupled with an amineterminated single-stranded DNA (ssDNA) oligonucleotide capture probe via a thiourea linkage. Confirmation of capture probe attachment was observed using fluorescent microscopy after hybridization of the capture probes with fluorescently labeled complimentary ssDNA oligonucleotides.

  9. New Concepts of Fluorescent Probes for Specific Detection of DNA Sequences: Bis-Modified Oligonucleotides in Excimer and Exciplex Detection

    Directory of Open Access Journals (Sweden)

    Gbaj A

    2009-01-01

    Full Text Available The detection of single base mismatches in DNA is important for diagnostics, treatment of genetic diseases, and identification of single nucleotide polymorphisms. Highly sensitive, specific assays are needed to investigate genetic samples from patients. The use of a simple fluorescent nucleoside analogue in detection of DNA sequence and point mutations by hybridisation in solution is described in this study. The 5’-bispyrene and 3’-naphthalene oligonucleotide probes form an exciplex on hybridisation to target in water and the 5’-bispyrene oligonucleotide alone is an adequate probe to determine concentration of target present. It was also indicated that this system has a potential to identify mismatches and insertions. The aim of this work was to investigate experimental structures and conditions that permit strong exciplex emission for nucleic acid detectors, and show how such exciplexes can register the presence of mismatches as required in SNP analysis. This study revealed that the hybridisation of 5'-bispyrenyl fluorophore to a DNA target results in formation of a fluorescent probe with high signal intensity change and specificity for detecting a complementary target in a homogeneous system. Detection of SNP mutations using this split-probe system is a highly specific, simple, and accessible method to meet the rigorous requirements of pharmacogenomic studies. Thus, it is possible for the system to act as SNP detectors and it shows promise for future applications in genetic testing.

  10. DNA-based stable isotope probing coupled with cultivation methods implicates Methylophaga in hydrocarbon degradation.

    Directory of Open Access Journals (Sweden)

    Sara eMishamandani

    2014-02-01

    Full Text Available Marine hydrocarbon-degrading bacteria perform a fundamental role in the oxidation and ultimate removal of crude oil and its petrochemical derivatives in coastal and open ocean environments. Those with an almost exclusive ability to utilize hydrocarbons as a sole carbon and energy source have been found confined to just a few genera. Here we used stable isotope probing (SIP, a valuable tool to link the phylogeny and function of targeted microbial groups, to investigate hydrocarbon-degrading bacteria in coastal North Carolina sea water (Beaufort Inlet, USA with uniformly labeled [13C]n-hexadecane. The dominant sequences in clone libraries constructed from 13C-enriched bacterial DNA (from n-hexadecane enrichments were identified to belong to the genus Alcanivorax, with ≤98% sequence identity to the closest type strain – thus representing a putative novel phylogenetic taxon within this genus. Unexpectedly, we also identified 13C-enriched sequences in heavy DNA fractions that were affiliated to the genus Methylophaga. This is a contentious group since, though some of its members have been proposed to degrade hydrocarbons, substantive evidence has not previously confirmed this. We used quantitative PCR primers targeting the 16S rRNA gene of the SIP-identified Alcanivorax and Methylophaga to determine their abundance in incubations amended with unlabeled n-hexadecane. Both showed substantial increases in gene copy number during the experiments. Subsequently, we isolated a strain representing the SIP-identified Methylophaga sequences (99.9% 16S rRNA gene sequence identity and used it to show, for the first time, direct evidence of hydrocarbon degradation by a cultured Methylophaga sp. This study demonstrates the value of coupling SIP with cultivation methods to identify and expand on the known diversity of hydrocarbon-degrading bacteria in the marine environment.

  11. Use of AffiProbe HPV test kit for detection of human papillomavirus DNA in genital scrapes.

    OpenAIRE

    Ranki, M; Leinonen, A W; Jalava, T. (Tiina); Nieminen, P.; Soares, V R; Paavonen, J; Kallio, A

    1990-01-01

    The presence of human papillomavirus (HPV) DNA in cervical and vaginal scrapes was analyzed by the AffiProbe HPV test kit (Orion Corp., Orion Pharmaceutica, Helsinki, Finland), which is a 1-day solution hybridization test for HPV type 6/11, 16, or 18. The AffiProbe test was compared with a commercially available dot blot test (ViraPap and ViraType tests; Life Technologies Inc., Gaithersburg, Md.). The study group consisted of 178 patients seen in a gynecological outpatient clinic. Altogether,...

  12. Cultivation-independent detection of autotrophic hydrogen-oxidizing bacteria by DNA stable-isotope probing.

    Science.gov (United States)

    Pumphrey, Graham M; Ranchou-Peyruse, Anthony; Spain, Jim C

    2011-07-01

    Knallgas bacteria are a physiologically defined group that is primarily studied using cultivation-dependent techniques. Given that current cultivation techniques fail to grow most bacteria, cultivation-independent techniques that selectively detect and identify knallgas bacteria will improve our ability to study their diversity and distribution. We used stable-isotope probing (SIP) to identify knallgas bacteria in rhizosphere soil of legumes and in a microbial mat from Obsidian Pool in Yellowstone National Park. When samples were incubated in the dark, incorporation of (13)CO(2) was H(2) dependent. SIP enabled the detection of knallgas bacteria that were not detected by cultivation, and the majority of bacteria identified in the rhizosphere soils were betaproteobacteria predominantly related to genera previously known to oxidize hydrogen. Bacteria in soil grew on hydrogen at concentrations as low as 100 ppm. A hydB homolog encoding a putative high-affinity NiFe hydrogenase was amplified from (13)C-labeled DNA from both vetch and clover rhizosphere soil. The results indicate that knallgas bacteria can be detected by SIP and populations that respond to different H(2) concentrations can be distinguished. The methods described here should be applicable to a variety of ecosystems and will enable the discovery of additional knallgas bacteria that are resistant to cultivation.

  13. The use of radionuclide DNA probe technology in epidemiological studies of leishmaniasis

    Energy Technology Data Exchange (ETDEWEB)

    Andrade, Antero Silva Ribeiro de [Centro de Desenvolvimento da Tecnologia Nuclear (CDTN), Belo Horizonte, MG (Brazil); Fernandes, Octavio [Fundacao Inst. Oswaldo Cruz (FIOCRUZ), Rio de Janeiro, RJ (Brazil). Dept. de Medicina Tropical; Gomes, Rosangela Fatima; Melo, Maria Norma de [Minas Gerais Univ., Belo Horizonte, MG (Brazil). Dept. de Parasitologia]|[Cor Jesus Fontes Mato Grosso Univ., Cuiaba, MT (Brazil). Hospital Universitario Julio Muller

    2000-07-01

    Cutaneous and mucosal leishmaniasis are due to different species that belongs to Leishmania (Leishmania) mexicana complex and Leishmania (Viannia) braziliensis complex, respectively. Cutaneous leishmaniasis ulcers may persist for months to years but eventually they heal, while mucocutaneous leishmaniasis can result in destructive lesions on the nose, oral pharynx, lips or face. The specific diagnosis of the disease is important because of the high cost and toxicity of the treatment and the subsequent medical follow-up depends on the precise identification of the complex that causes the infection. The epidemiological information furnished by the identification of parasites of a given endemic region is also essential for the design of appropriate control measures. In this work we typed, using specific DNA probes labelled with {sup 32}P radionuclide, samples collected from patients living in endemic areas of Mato Grosso state. The results showed that L. braziliensis is the predominant group infecting human patients in the state. We have typed, up to the moment 68 samples. 64 samples (94.1%) belonged to the L. braziliensis complex and only 4 (5.9%) belonged to the L. mexicana complex. (author)

  14. Multiple DNA Extractions Coupled with Stable-Isotope Probing of Anthracene-Degrading Bacteria in Contaminated Soil▿†

    Science.gov (United States)

    Jones, Maiysha D.; Singleton, David R.; Sun, Wei; Aitken, Michael D.

    2011-01-01

    In many of the DNA-based stable-isotope probing (SIP) studies published to date in which soil communities were investigated, a single DNA extraction was performed on the soil sample, usually using a commercial DNA extraction kit, prior to recovering the 13C-labeled (heavy) DNA by density-gradient ultracentrifugation. Recent evidence suggests, however, that a single extraction of a soil sample may not lead to representative recovery of DNA from all of the organisms in the sample. To determine whether multiple DNA extractions would affect the DNA yield, the eubacterial 16S rRNA gene copy number, or the identification of anthracene-degrading bacteria, we performed seven successive DNA extractions on the same aliquot of contaminated soil either untreated or enriched with [U-13C]anthracene. Multiple extractions were necessary to maximize the DNA yield and 16S rRNA gene copy number from both untreated and anthracene-enriched soil samples. Sequences within the order Sphingomonadales, but unrelated to any previously described genus, dominated the 16S rRNA gene clone libraries derived from 13C-enriched DNA and were designated “anthracene group 1.” Sequences clustering with Variovorax spp., which were also highly represented, and sequences related to the genus Pigmentiphaga were newly associated with anthracene degradation. The bacterial groups collectively identified across all seven extracts were all recovered in the first extract, although quantitative PCR analysis of SIP-identified groups revealed quantitative differences in extraction patterns. These results suggest that performing multiple DNA extractions on soil samples improves the extractable DNA yield and the number of quantifiable eubacterial 16S rRNA gene copies but have little qualitative effect on the identification of the bacterial groups associated with the degradation of a given carbon source by SIP. PMID:21398486

  15. Multiple DNA extractions coupled with stable-isotope probing of anthracene-degrading bacteria in contaminated soil.

    Science.gov (United States)

    Jones, Maiysha D; Singleton, David R; Sun, Wei; Aitken, Michael D

    2011-05-01

    In many of the DNA-based stable-isotope probing (SIP) studies published to date in which soil communities were investigated, a single DNA extraction was performed on the soil sample, usually using a commercial DNA extraction kit, prior to recovering the (13)C-labeled (heavy) DNA by density-gradient ultracentrifugation. Recent evidence suggests, however, that a single extraction of a soil sample may not lead to representative recovery of DNA from all of the organisms in the sample. To determine whether multiple DNA extractions would affect the DNA yield, the eubacterial 16S rRNA gene copy number, or the identification of anthracene-degrading bacteria, we performed seven successive DNA extractions on the same aliquot of contaminated soil either untreated or enriched with [U-(13)C]anthracene. Multiple extractions were necessary to maximize the DNA yield and 16S rRNA gene copy number from both untreated and anthracene-enriched soil samples. Sequences within the order Sphingomonadales, but unrelated to any previously described genus, dominated the 16S rRNA gene clone libraries derived from (13)C-enriched DNA and were designated "anthracene group 1." Sequences clustering with Variovorax spp., which were also highly represented, and sequences related to the genus Pigmentiphaga were newly associated with anthracene degradation. The bacterial groups collectively identified across all seven extracts were all recovered in the first extract, although quantitative PCR analysis of SIP-identified groups revealed quantitative differences in extraction patterns. These results suggest that performing multiple DNA extractions on soil samples improves the extractable DNA yield and the number of quantifiable eubacterial 16S rRNA gene copies but have little qualitative effect on the identification of the bacterial groups associated with the degradation of a given carbon source by SIP.

  16. Fiber-optic communication systems

    CERN Document Server

    Agrawal, Govind P

    2010-01-01

    This book provides a comprehensive account of fiber-optic communication systems. The 3rd edition of this book is used worldwide as a textbook in many universities. This 4th edition incorporates recent advances that have occurred, in particular two new chapters. One deals with the advanced modulation formats (such as DPSK, QPSK, and QAM) that are increasingly being used for improving spectral efficiency of WDM lightwave systems. The second chapter focuses on new techniques such as all-optical regeneration that are under development and likely to be used in future communication systems. All othe

  17. Fetal sex determination in the first trimester of pregnancy using a Y chromosome-specific DNA probe

    Energy Technology Data Exchange (ETDEWEB)

    Zeng, Y.; Huang, S.; Chen, M.; Huang, Y.; Zhang, M.; Dong, J.; Ku, A.; Xu, S.

    1987-05-01

    Prenatal determination of fetal sex is important for the prevention of X-linked disorders such as hemophilia, Lesch-Nyhan syndrome and Duchenne muscular dystrophy. The complex procedures of prenatal diagnosis for X-linked disorders are unnecessary if the fetus is female, because usually no clinical symptoms ever appear in female. pY 3.4 probe used in this work for sex determination is a 3.4 kilobase human repeat sequence. The probe is specific for the Y chromosome of males and can be used for sex determination. The other prove pBLUR used in this paper as control is a widely dispersed, highly repeated human Alu family DNA sequence, represented equally in male and female DNA. On the basis of the relative densities of the autoradiographic spots produced by hybridization of fetal DNA with pY3.4 and pBLUR, the sex of fetus can be clearly identified. Further the authors can determine the radioactive intensity (cpm) of the hybridized DNA spots and the ratio of hybridization with Y3.4 to pBLUR (Y3.4/pBLUR x 10). Results show that the hybridization ratio of DNA from chorionic villi of male (1.03 +/- 0.24) is significantly higher than that of female (0.16 +/- 0.09). Therefore, sex determination of the fetus can be made, based on the ratio of pY3.4/pBLUR x 10. If necessary they can also use Southern hybridization with pY 3.4 probe of DNA isolated from chorionic villi to confirm the result of dot hybridization.

  18. Supersandwich cytosensor for selective and ultrasensitive detection of cancer cells using aptamer-DNA concatamer-quantum dots probes.

    Science.gov (United States)

    Liu, Hongying; Xu, Shouming; He, Zhimei; Deng, Anping; Zhu, Jun-Jie

    2013-03-19

    In this work, a signal amplification supersandwich strategy was developed for highly selective and sensitive detection of cancer cells using aptamer-DNA concatamer-quantum dots (QDs) probes. First of all, electrode materials denoted as MWCNTs@PDA@AuNPs were fabricated by multiwall carbon nanotubes (MWCNTs), gold nanoparticles (AuNPs), and polydopamine (PDA) using a layer-by-layer technique. Then, the prepared bases as matrices were applied to bind concanavalin A (Con A), resulting in high stability, bioactivity, and capability for cell capture. Meanwhile, aptamer-DNA concatamer-QDs were designed via DNA hybridization followed by covalent assembling, which incorporated the specific recognition of the aptamer with the signal amplification of the DNA concatamer and QDs. With aptamer-DNA concatamer-QDs as recognizing probes, the model cancer cells (CCRF-CEM cells) were detected using a MWCNTs@PDA@AuNPs modified electrode with trapped Con A by means of fluorescence and electrochemical methods. The proposed supersandwich cytosensor showed high sensitivity with the detection limit of 50 cells mL(-1). More importantly, it could distinguish cancer cells from normal cells, which indicated the promising applications of our method in clinical diagnosis and treatment of cancers.

  19. Synthesis of novel fluorescent probe Tb(III)-7-carboxymethoxy-4-methylcoumarin complex for sensing of DNA

    Energy Technology Data Exchange (ETDEWEB)

    Hussein, Belal H.M., E-mail: belalhussein102@yahoo.com [Department of Chemistry, Faculty of Science, Suez Canal University, Ismailia (Egypt); Azab, Hassan A. [Department of Chemistry, Faculty of Science, Suez Canal University, Ismailia (Egypt); Fathalla, Walid [Department of Mathematical and Physical Sciences, Faculty of Engineering, Port-Said University, Port-Said (Egypt); Ali, Sherin A.M. [Department of Mathematical and Physical Sciences, Faculty of Engineering, Suez Canal University, Ismailia (Egypt)

    2013-02-15

    New fluorescent probe Tb(III) (7-carboxymethoxy-4-methylcoumarin)2(SCN) (C2H5OH)(H2O) was synthesized and characterized by spectroscopy and thermal analysis. The absorption and fluorescence spectra of 7-carboxymethoxy-4-methylcoumarin (CMMC) and Tb(III)-CMMC complex have been measured in different solvents. The interactions of Tb(III)-CMMC complex with calf thymus nucleic acid (CT-DNA) have been investigated using steady state fluorescence measurements. The changes in the fluorescence intensity have been used for the quantitative determination of DNA with LOD of 3.45 ng in methanol-water (9:1, v/v). The association constants of DNA with Tb(III)-CMMC complex was found to be 2.62 Multiplication-Sign 1010 M{sup -1}. - Highlights: Black-Right-Pointing-Pointer New fluorescent probe Terbium (III)-7-carboxy methoxy-4-methylcoumarin complex has been synthesized and characterized. Black-Right-Pointing-Pointer FTIR spectrum of Tb(III)-complex shows a characteristic band for thiocyanate group. Black-Right-Pointing-Pointer DNA interaction with Terbium (III)-7-carboxy methoxy-4-methylcoumarin has been studied by fluorescence techniques. Black-Right-Pointing-Pointer The change in the fluorescence intensity has been used for the quantitative determination of DNA. Black-Right-Pointing-Pointer The result was better than most of the well-known methods including the ethidium bromide method.

  20. Method to detect the end-point for PCR DNA amplification using an ionically labeled probe and measuring impedance change

    Science.gov (United States)

    Miles, Robin R.; Belgrader, Phillip; Fuller, Christopher D.

    2007-01-02

    Impedance measurements are used to detect the end-point for PCR DNA amplification. A pair of spaced electrodes are located on a surface of a microfluidic channel and an AC or DC voltage is applied across the electrodes to produce an electric field. An ionically labeled probe will attach to a complementary DNA segment, and a polymerase enzyme will release the ionic label. This causes the conductivity of the solution in the area of the electrode to change. This change in conductivity is measured as a change in the impedance been the two electrodes.

  1. Electrochemical DNA probe for Hg(2+) detection based on a triple-helix DNA and Multistage Signal Amplification Strategy.

    Science.gov (United States)

    Wang, Huan; Zhang, Yihe; Ma, Hongmin; Ren, Xiang; Wang, Yaoguang; Zhang, Yong; Wei, Qin

    2016-12-15

    In this work, an ultrasensitive electrochemical sensor was developed for detection of Hg(2+). Gold nanoparticles decorated bovine serum albumin reduction of graphene oxide (AuNP-BSA-rGO) were used as subsurface material for the immobilization of triple-helix DNA. The triple-helix DNA containing a thiol labelled single-stranded DNA (sDNA) and a thymine-rich DNA (T-rich DNA), which could be unwinded in the present of Hg(2+) to form more stable thymine-Hg(2+)-thymine (T-Hg(2+)-T) complex. T-Hg(2+)-T complex was then removed and the sDNA was left on the electrode. At this time, gold nanoparticle carrying thiol labelled cytosine-rich complementary DNA (cDNA-AuNP) could bind with the free sDNA. Meanwhile, the other free cDNA on AuNP could bind with each other in the present of Ag(+) to form the stable cytosine-Ag(+)-cytosine (C-Ag(+)-C) complex and circle amplification. Plenty of C-Ag(+)-C could form silver nanoclusters by electrochemical reduction and the striping signal of Ag could be measured for purpose of the final electrochemical detection of Hg(2+). This sensor could detect Hg(2+) over a wide concentration range from 0.1 to 130nM with a detection limit of 0.03nM.

  2. Interfacing click chemistry with automated oligonucleotide synthesis for the preparation of fluorescent DNA probes containing internal xanthene and cyanine dyes.

    Science.gov (United States)

    Astakhova, I Kira; Wengel, Jesper

    2013-01-14

    Double-labeled oligonucleotide probes containing fluorophores interacting by energy-transfer mechanisms are essential for modern bioanalysis, molecular diagnostics, and in vivo imaging techniques. Although bright xanthene and cyanine dyes are gaining increased prominence within these fields, little attention has thus far been paid to probes containing these dyes internally attached, a fact which is mainly due to the quite challenging synthesis of such oligonucleotide probes. Herein, by using 2'-O-propargyl uridine phosphoramidite and a series of xanthenes and cyanine azide derivatives, we have for the first time performed solid-phase copper(I)-catalyzed azide-alkyne cycloaddition (CuAAC) click labeling during the automated phosphoramidite oligonucleotide synthesis followed by postsynthetic click reactions in solution. We demonstrate that our novel strategy is rapid and efficient for the preparation of novel oligonucleotide probes containing internally positioned xanthene and cyanine dye pairs and thus represents a significant step forward for the preparation of advanced fluorescent oligonucleotide probes. Furthermore, we demonstrate that the novel xanthene and cyanine labeled probes display unusual and very promising photophysical properties resulting from energy-transfer interactions between the fluorophores controlled by nucleic acid assembly. Potential benefits of using these novel fluorescent probes within, for example, molecular diagnostics and fluorescence microscopy include: Considerable Stokes shifts (40-110 nm), quenched fluorescence of single-stranded probes accompanied by up to 7.7-fold light-up effect of emission upon target DNA/RNA binding, remarkable sensitivity to single-nucleotide mismatches, generally high fluorescence brightness values (FB up to 26), and hence low limit of target detection values (LOD down to <5 nM).

  3. The use of radionuclide DNA probe technology for epidemiological studies of tegumentary leishmaniasis in Mato Grosso state, Brazil

    Energy Technology Data Exchange (ETDEWEB)

    Andrade, Antero Silva Ribeiro de [Centro de Desenvolvimento da Tecnologia Nuclear (CDTN/CNEN), Belo Horizonte, MG (Brazil); Fernandes, Octavio [Fundacao Oswaldo Cruz (FIOCRUZ), Rio de Janeiro, RJ (Brazil). Dept. de Medicina Tropical; Heub, Marcia; Fontes, Cor Jesus [Universidade Federal do Mato Grosso, Cuiaba, MT (Brazil). Hospital Universitario Julio Muller; Carvalho, Maria de Lourdes Ribeiro; Melo, Maria Norma de [Universidade Federal de Minas Gerais, Belo Horizonte, MG (Brazil). Dept. de Parasitologia

    2005-10-15

    DNA hybridisation, using probes labelled with 32 P, was used to type Leishmania samples isolated from patients living in endemic areas of Mato Grosso State (Brazil), and clinically diagnosed as having tegumentary leishmaniasis. k DNA cloned mini-circle probes specific for the Leishmania mexicana and Leishmania braziliensis complexes were used. The results showed that L. braziliensis is the predominant group infecting human patients in the state. Sixty-eight samples were typed, 64 samples (94.1%) belonging to the L. braziliensis complex and only four (5.9%) belonging to the L. mexicana complex. Accurate identification of the Leishmania permits better orientation of the medical follow-up, since clinical manifestations may vary depending on the complex to which the parasite belongs. The epidemiological information furnished by the identification of the Leishmania in given endemic area is also essential for the design of appropriate control measures. (author)

  4. Use of a multi-thermal washer for DNA microarrays simplifies probe design and gives robust genotyping assays

    DEFF Research Database (Denmark)

    Petersen, J.; Poulsen, Lena; Petronis, S.

    2008-01-01

    DNA microarrays are generally operated at a single condition, which severely limits the freedom of designing probes for allele-specific hybridization assays. Here, we demonstrate a fluidic device for multi-stringency posthybridization washing of microarrays on microscope slides. This device...... is called a multi-thermal array washer (MTAW), and it has eight individually controlled heating zones, each of which corresponds to the location of a subarray on a slide. Allele-specific oligonucleotide probes for nine mutations in the beta-globin gene were spotted in eight identical subarrays at positions...... corresponding to the temperature zones of the MTAW. After hybridization with amplified patient material, the slides were mounted in the MTAW, and each subarray was exposed to different temperatures ranging from 22 to 40 degrees C. When processed in the MTAW, probes selected without considering melting...

  5. Clinical utility of a DNA probe to 17p11.2 in screening of patients with a peripheral neuropathy

    Energy Technology Data Exchange (ETDEWEB)

    Blancato, J.; Precht, K.; Meck, J. [Georgetown Univ., Hospital, Washington, DC (United States)] [and others

    1994-09-01

    We assessed the usefulness of in situ hybridization with a DNA probe to the area of chromosome 17 at p11.2 as a diagnostic tool for screening for Charcot Marte Tooth 1A (CMT 1A). In situ hybridization with a probe to 17p11.2 was performed on fixed lymphocytes from the following groups of individuals: (1) normal controls; (2) patients evoking a strong clinical suspicion of CMT 1A; and (3) 3 families with an apparent autosomal dominant peripheral neuropathy of unknown diagnoses. Group 2 patients had evidence of demyelination as defined by nerve conduction of less that 50% of the normal mean or terminal latency greater than 50% of the normal mean in conduction studies. Analysis of interphase cells hybridized with a cosmid DNA probe to 17p11.2 requires inclusion of a normal control with each trial and masked observer. Due to the size of the target DNA and the nature of the centromeric heterochromatin, the scoring of this probe is more subjective than centromere probes. For example, if the two 17 chromosomes are decondensed as in interphase, two tandem signals may be visualized as one. Results from duplication positive patients demonstrate a large proportion of cells with two closely aligned, but separate, signals with an additional single signal. Normal results demonstrate a majority of cells with two separate signals representing both normal homologues. None of the 3 families with questionable diagnosis revealed a duplication at the region, reinforcing our belief that a clinical diagnosis is the most discriminating tool available for diagnosis of CMT 1A. We concur with Boylan that molecular analysis for CMT 1A is useful for establishing a diagnosis of CMT 1A, but is not a primary differential diagnostic test. The yield in screening patients without physiologic evidence of demyelination is likely to be low. We further find that the use of in situ hybridization is a simple method of performing the duplication analysis.

  6. Target-induced reconfiguration of DNA probes for recycling amplification and signal-on electrochemical detection of hereditary tyrosinemia type I gene.

    Science.gov (United States)

    Dou, Baoting; Yang, Cuiyun; Chai, Yaqin; Yuan, Ruo; Xiang, Yun

    2015-09-01

    By coupling target DNA-induced reconfiguration of the dsDNA probes with enzyme-assisted target recycling amplification, we describe the development of a signal-on electrochemical sensing approach for sensitive detection of hereditary tyrosinemia type I gene. The dsDNA probes are self-assembled on the sensing electrode, and the addition of the target DNA reconfigures and switches the dsDNA probes into active substrates for exonuclease III, which catalytically digests the probes and leads to cyclic reuse of the target DNA. The target DNA recycling and the removal of one of the ssDNA from the dsDNA probes by exonuclease III result in the formation of many hairpin structures on the sensor surface, which brings the electroactive methylene blue labels into proximity with the electrode and produces a significantly amplified current response for sensitive detection of the target gene down to 0.24 pM. This method is also selective to discriminate single-base mismatch and can be employed to detect the target gene in human serum samples. With the demonstration for the detection of the target gene, we expect the developed method to be a universal sensitive sensing platform for the detection of different nucleic acid sequences.

  7. Probing DNA-stabilized fluorescent silver nanocluster spectral heterogeneity by time-correlated single photon counting

    DEFF Research Database (Denmark)

    Carro, Miguel; Paolucci, Valentina; Hooley, Emma Nicole

    2016-01-01

    DNA-stabilized silver nanoclusters (DNA-AgNCs) are promising fluorophores whose photophysical properties and synthesis procedures have received increased attention in the literature. However, depending on the preparation conditions and the DNA sequence, the DNA-AgNC samples can host a range of di...

  8. A comparative cytogenetic study of Drosophila parasitoids (Hymenoptera, Figitidae) using DNA-binding fluorochromes and FISH with 45S rDNA probe.

    Science.gov (United States)

    Gokhman, Vladimir E; Bolsheva, Nadezhda L; Govind, Shubha; Muravenko, Olga V

    2016-06-01

    Karyotypes of Leptopilina boulardi (Barbotin, Carton et Keiner-Pillault, 1979) (n = 9), L. heterotoma (Thomson, 1862) (n = 10), L. victoriae Nordlander, 1980 (n = 10) and Ganaspis xanthopoda (Ashmead, 1896) (n = 9) (Hymenoptera, Figitidae) were studied using DNA-binding ligands with different base specificity [propidium iodide (PI), chromomycin A3 (CMA3) and 4',6-diamidino-2-phenylindole (DAPI)], and fluorescence in situ hybridization (FISH) with a 45S rDNA probe. Fluorochrome staining was similar between the different fluorochromes, except for a single CMA3- and PI-positive and DAPI-negative band per haploid karyotype of each species. FISH with 45S rDNA probe detected a single rDNA site in place of the bright CMA3-positive band, thus identifying the nucleolus organizing region (NOR). Chromosomal locations of NORs were similar for both L. heterotoma and L. victoriae, but strongly differed in L. boulardi as well as in G. xanthopoda. Phylogenetic aspects of NOR localization in all studied species are briefly discussed.

  9. Rapid quantification of hepatitis B virus DNA by real-time PCR using efficient TaqMan probe and extraction of virus DNA

    Institute of Scientific and Technical Information of China (English)

    Yan-Qin Lu; Jin-Xiang Han; Peng Qi; Wei Xu; Yan-Hui Zu; Bo Zhu

    2006-01-01

    AIM: To rapidly quantify hepatitis B virus (HBV) DNA by real-time PCR using efficient TaqMan probe and extraction methods of virus DNA.METHODS: Three standards were prepared by cloning PCR products which targeted S, C and X region of HBV genome into pGEM-T vector respectively. A pair of primers and matched TaqMan probe were selected by comparing the copy number and the Ct values of HBV serum samples derived from the three different standard curves using certain serum DNA. Then the efficiency of six HBV DNA extraction methods including guanidinium isothiocyanate, proteinase K, NaI, NaOH lysis, alkaline lysis and simple boiling was analyzed in sample A, B and C by real-time PCR. Meanwhile, 8 clinical HBV serum samples were quantified.RESULTS: The copy number of the same HBV serum sample originated from the standard curve of S, C and Xregions was 5.7 x 104/mL, 6.3 x 102/mL and 1.6 x 103/mL respectively. The relative Ct value was 26.6, 31.8 and 29.5 respectively. Therefore, primers and matched probe from S region were chosen for further optimization of six extraction methods. The copy number of HBV serum samples A, B and C was 3.49 x 109/mL, 2.08 x 106/mL and 4.40 x 107/mL respectively, the relative Ct value was 19.9, 30 and 26.2 in the method of NaOH lysis,which was the efficientest among six methods. Simple boiling showed a slightly lower efficiency than NaOH lysis. Guanidinium isothiocyanate, proteinase K and NaI displayed that the copy number of HBV serum sample A,B and C was around 105/mL, meanwhile the Ct value was about 30. Alkaline failed to quantify the copy number of three HBV serum samples. Standard deviation (SD) and coefficient variation (CV) were very low in all 8 clinical HBV serum samples, showing that quantification of HBV DNA in triplicate was reliable and accurate.CONCLUSION: Real-time PCR based on optimized primers and TaqMan probe from S region in combination with NaOH lysis is a simple, rapid and accurate method for quantification of HBV serum DNA.

  10. Detection of mRNA of the cyclin D1 breast cancer marker by a novel duplex-DNA probe.

    Science.gov (United States)

    Segal, Meirav; Yavin, Eylon; Kafri, Pinhas; Shav-Tal, Yaron; Fischer, Bilha

    2013-06-27

    Previously, we have described 5-((4-methoxy-phenyl)-trans-vinyl)-2'-deoxy-uridine, 6, as a fluorescent uridine analogue exhibiting a 3000-fold higher quantum yield (Φ 0.12) and maximum emission (478 nm) which is 170 nm red-shifted as compared to uridine. Here, we utilized 6 for preparation of labeled oligodeoxynucleotide (ODN) probes based on MS2 and cyclin D1 (a known breast cancer mRNA marker) sequences. Cyclin D1-derived labeled-ssODN showed a 9.5-fold decrease of quantum yield upon duplex formation. On the basis of this finding, we developed the ds-NIF (nucleoside with intrinsic fluorescence)-probe methodology for detection of cyclin D1 mRNA, by which the fluorescent probe is released upon recognition of target mRNA by the relatively dark NIF-duplex-probe. Indeed, we successfully detected, a ss-deoxynucleic acid (DNA) variant of cyclin D1 mRNA using a dark NIF-labeled duplex-probe, and monitoring the recognition process by fluorescence spectroscopy and gel electrophoresis. Furthermore, we successfully detected cyclin D1 mRNA in RNA extracted from cancerous human cells, using ds-NIF methodology.

  11. Development of Fluorescent Protein Probes Specific for Parallel DNA and RNA G-Quadruplexes.

    Science.gov (United States)

    Dang, Dung Thanh; Phan, Anh Tuân

    2016-01-01

    We have developed fluorescent protein probes specific for parallel G-quadruplexes by attaching cyan fluorescent protein to the G-quadruplex-binding motif of the RNA helicase RHAU. Fluorescent probes containing RHAU peptide fragments of different lengths were constructed, and their binding to G-quadruplexes was characterized. The selective recognition and discrimination of G-quadruplex topologies by the fluorescent protein probes was easily detected by the naked eye or by conventional gel imaging.

  12. Impedimetric genosensor for detection of hepatitis C virus (HCV1) DNA using viral probe on methylene blue doped silica nanoparticles.

    Science.gov (United States)

    Singhal, Chaitali; Ingle, Aviraj; Chakraborty, Dhritiman; Pn, Anoop Krishna; Pundir, C S; Narang, Jagriti

    2017-05-01

    An impedimetric genosensor was fabricated for detection of hepatitis C virus (HCV) genotype 1 in serum, based on hybridization of the probe with complementary target cDNA from sample. To achieve it, probe DNA complementary to HCVgene was immobilized on the surface of methylene blue (MB) doped silica nanoparticles MB@SiNPs) modified fluorine doped tin oxide (FTO) electrode. The synthesized MB@SiNPs was characterized using scanning electron microscopy (SEM), high resolution transmission electron microscopy (HRTEM) and X-ray diffraction (XRD) pattern. This modified electrode (ssDNA/MB@SiNPs/FTO) served both as a signal amplification platform (due to silica nanoparticles (SiNPs) as well as an electrochemical indicator (due to methylene blue (MB)) for the detection of the HCV DNA in patient serum sample. The genosensor was optimized and evaluated. The sensor showed a dynamic linear range 100-10(6) copies/mL, with a detection limit of 90 copies/mL. The sensor was applied for detection of HCV in sera of hepatitis patient and could be renewed. The half life of the sensor was 4 weeks. The MB@SiNPs/FTO electrode could be used for preparation of other gensensors also. Copyright © 2017 Elsevier B.V. All rights reserved.

  13. Micron-sized surface enhanced Raman scattering reporter/fluorescence probe encoded colloidal microspheres for sensitive DNA detection.

    Science.gov (United States)

    You, Lijun; Li, Ruimin; Dong, Xu; Wang, Fang; Guo, Jia; Wang, Changchun

    2017-02-15

    A new type of optical probes, featuring surface enhanced Raman scattering (SERS) and fluorescence spectra dual-mode encoding, has been reported in this article. Based on the uniform micrometer-sized melamine resin/Ag nanoparticles (MRM/Ag-NPs) composite microspheres, the SERS reporters and fluorescent probes were successfully fixed onto the different layers of the MEM/Ag-NPs microspheres, which supported the sensitive DNA detecton. The two spectroscopic methods commonly considered to be contradictive to each other, yet the optical signals were separable in the experiments. The dual-encoding strategy and single microsphere detecton method put the number of available independent codes to be rough the multiple of those available in the two optical detection channels, which increases far more rapidly than the summation of the two channels. As a proof of cencept, the utility of this dual spectrum mode SERS-fluoresecence encoded microsphere (SFEM) was demonstrated in a specific DNA detection using complimentary ssDNA functionalized magnetic beads as the DNA capturing and separation agents. Excellent encoding results were demonstrated from the decoding of the SERS and fluorescence signals of the SFEM. The method appears to be general in scope and we expect that the SERS-fluoresecence encoded microspheres system is applicable to multiplex bioassays of a variety of biomolecules.

  14. A combined approach of DNA probe and RFLP for family and species identification of larval stages of commercially important aquatic species: A study on the surfclam Spisula solidissima

    Digital Repository Service at National Institute of Oceanography (India)

    Achuthankutty, C.T.

    This paper deals briefly with a technique developed for the identification of the early stages of veligers of surfclam Spisula solidissima from the larval stages of other common bivalve species using a combination of DNA probe and restriction...

  15. Surface plasmon resonator using high sensitive resonance telecommunication wavelengths for DNA sensors of Mycobacterium tuberculosis with thiol-modified probes.

    Science.gov (United States)

    Hsu, Shih-Hsiang; Hung, Shao-Chiang; Chen, Yu-Kun; Jian, Zhi-Hao

    2014-12-25

    Various analytes can be verified by surface plasmon resonance, thus continuous improvement of this sensing technology is crucial for better sensing selection and higher sensitivity. The SPR sensitivity on the wavelength modulation is enhanced with increasing wavelengths. The telecommunication wavelength range was then utilized to detect Mycobacterium tuberculosis (MTB) deoxyribonucleic acid (DNA) under two situations, without immobilization and with 5'-thiol end labeled IS6100 DNA probes, for SPR sensitivity comparison. The experimental data demonstrated that the SPR sensitivity increased more than 13 times with the wavelength modulation after immobilization. Since the operating wavelength accuracy of a tunable laser source can be controlled within 0.001 nm, the sensitivity and resolution on immobilized MTB DNA were determined as 1.04 nm/(μg/mL) and 0.9 ng/mL, respectively.

  16. Transcription-factor-mediated DNA looping probed by high-resolution, single-molecule imaging in live E. coli cells.

    Science.gov (United States)

    Hensel, Zach; Weng, Xiaoli; Lagda, Arvin Cesar; Xiao, Jie

    2013-01-01

    DNA looping mediated by transcription factors plays critical roles in prokaryotic gene regulation. The "genetic switch" of bacteriophage λ determines whether a prophage stays incorporated in the E. coli chromosome or enters the lytic cycle of phage propagation and cell lysis. Past studies have shown that long-range DNA interactions between the operator sequences O(R) and O(L) (separated by 2.3 kb), mediated by the λ repressor CI (accession number P03034), play key roles in regulating the λ switch. In vitro, it was demonstrated that DNA segments harboring the operator sequences formed loops in the presence of CI, but CI-mediated DNA looping has not been directly visualized in vivo, hindering a deep understanding of the corresponding dynamics in realistic cellular environments. We report a high-resolution, single-molecule imaging method to probe CI-mediated DNA looping in live E. coli cells. We labeled two DNA loci with differently colored fluorescent fusion proteins and tracked their separations in real time with ∼40 nm accuracy, enabling the first direct analysis of transcription-factor-mediated DNA looping in live cells. Combining looping measurements with measurements of CI expression levels in different operator mutants, we show quantitatively that DNA looping activates transcription and enhances repression. Further, we estimated the upper bound of the rate of conformational change from the unlooped to the looped state, and discuss how chromosome compaction may impact looping kinetics. Our results provide insights into transcription-factor-mediated DNA looping in a variety of operator and CI mutant backgrounds in vivo, and our methodology can be applied to a broad range of questions regarding chromosome conformations in prokaryotes and higher organisms.

  17. Transcription-factor-mediated DNA looping probed by high-resolution, single-molecule imaging in live E. coli cells.

    Directory of Open Access Journals (Sweden)

    Zach Hensel

    Full Text Available DNA looping mediated by transcription factors plays critical roles in prokaryotic gene regulation. The "genetic switch" of bacteriophage λ determines whether a prophage stays incorporated in the E. coli chromosome or enters the lytic cycle of phage propagation and cell lysis. Past studies have shown that long-range DNA interactions between the operator sequences O(R and O(L (separated by 2.3 kb, mediated by the λ repressor CI (accession number P03034, play key roles in regulating the λ switch. In vitro, it was demonstrated that DNA segments harboring the operator sequences formed loops in the presence of CI, but CI-mediated DNA looping has not been directly visualized in vivo, hindering a deep understanding of the corresponding dynamics in realistic cellular environments. We report a high-resolution, single-molecule imaging method to probe CI-mediated DNA looping in live E. coli cells. We labeled two DNA loci with differently colored fluorescent fusion proteins and tracked their separations in real time with ∼40 nm accuracy, enabling the first direct analysis of transcription-factor-mediated DNA looping in live cells. Combining looping measurements with measurements of CI expression levels in different operator mutants, we show quantitatively that DNA looping activates transcription and enhances repression. Further, we estimated the upper bound of the rate of conformational change from the unlooped to the looped state, and discuss how chromosome compaction may impact looping kinetics. Our results provide insights into transcription-factor-mediated DNA looping in a variety of operator and CI mutant backgrounds in vivo, and our methodology can be applied to a broad range of questions regarding chromosome conformations in prokaryotes and higher organisms.

  18. Kinetic adsorption profile and conformation evolution at the DNA-gold nanoparticle interface probed by dynamic light scattering.

    Science.gov (United States)

    Wang, Wenjie; Ding, XiaoFan; He, Miao; Wang, Jing; Lou, Xinhui

    2014-10-21

    The kinetic adsorption profile at the DNA-gold nanoparticle (AuNP) interface is probed by following the binding and organization of thiolated linear DNA and aptamers of varying chain lengths (15, 30, 44, and 51 mer) to the surface of AuNPs (13.0 ± 1.0 nm diameter). A systematic investigation utilizing dynamic light scattering has been performed to directly measure the changes in particle size during the course of a typical aging-salting thiolated DNA/AuNP preparation procedure. We discuss the effect of DNA chain length, composition, salt concentration, and secondary structure on the kinetics and conformation at the DNA-AuNP interface. The adsorption kinetics are chain-length dependent, composition independent, and not diffusion rate limited for the conditions we report here. The kinetic data support a mechanism of stepwise adsorption of thiols to the surface of AuNPs and reorganization of the thiols at the interface. Very interestingly, the kinetic increases of the particle sizes are modeled accurately by the pseudo-second-order rate model, suggesting that DNA could possess the statistically well-defined conformational evolution. Together with other experimental evidence, we propose a dynamic inner-layer and outer-tail (DILOT) model to describe the evolution of the DNA conformation after the initial adsorption of a single oligonucleotide layer. According to this model, the length of the tails that extend from the surface of AuNPs, capable for hybridization or molecular recognition, can be conveniently calculated. Considering the wide applications of DNA/AuNPs, the results should have important implications in sensing and DNA-directed nanoparticle assembly.

  19. DNA interstrand cross-links of an antitumor trinuclear platinum(II) complex: thermodynamic analysis and chemical probing.

    Science.gov (United States)

    Malina, Jaroslav; Farrell, Nicholas P; Brabec, Viktor

    2011-06-06

    The trinuclear platinum compound [{trans-PtCl(NH(3))(2)}(2)(μ-trans-Pt(NH(3))(2){NH(2)(CH(2))(6)NH(2)}(2))](4+) (BBR3464) belongs to the polynuclear class of platinum-based anticancer agents. These agents form in DNA long-range (Pt,Pt) interstrand cross-links, whose role in the antitumor effects of BBR3464 predominates. Our results show for the first time that the interstrand cross-links formed by BBR3464 between two guanine bases in opposite strands separated by two base pairs (1,4-interstrand cross-links) exist as two distinct conformers, which are not interconvertible, not only if these cross-links are formed in the 5'-5', but also in the less-usual 3'-3' direction. Analysis of the conformers by differential scanning calorimetry, chemical probes of DNA conformation, and minor groove binder Hoechst 33258 demonstrate that each of the four conformers affects DNA in a distinctly different way and adopts a different conformation. The results also support the thesis that the molecule of antitumor BBR3464 when forming DNA interstrand cross-links may adopt different global structures, including different configurations of the linker chain of BBR3464 in the minor groove of DNA. Our findings suggest that the multiple DNA interstrand cross-links available to BBR3464 may all contribute substantially to its cytotoxicity.

  20. Advanced Components For Fiber-Optical Systems

    Science.gov (United States)

    Depaula, Ramon; Stowe, David W.

    1989-01-01

    Paper reviews statuses of some advanced passive and active optical components for use with optical fibers. Emphasis on highly birefringent components controling polarization, because control of polarization critical in applications as fiber-optical gyroscopes, interferometric sensors, and coherent communications.

  1. A DNA fingerprint probe from Mycosphaerella graminicola identifies an active transposable element

    NARCIS (Netherlands)

    Goodwin, S.B.; Cavaletto, J.R.; Waalwijk, C.; Kema, G.H.J.

    2001-01-01

    DNA fingerprinting has been used extensively to characterize populations of Mycosphaerella graminicola, the Septoria tritici blotch pathogen of wheat. The highly polymorphic DNA fingerprints of Mycosphaerella graminicola were assumed to reflect the action of transposable elements. However, there was

  2. DNA Probe for Identification of the Take-All Fungus, Gaeumannomyces graminis.

    Science.gov (United States)

    Henson, J M

    1989-02-01

    A 4.3-kilobase mitochondrial DNA fragment was cloned from Gaeumannomyces graminis var. tritici, the causative agent of take-all disease of wheat. Although this DNA fragment hybridized with all three varieties of G. graminis, it showed little homology with DNA from other fungi and thus should be useful for identification of Gaeumannomyces sp. recovered from infected plants.

  3. DNA Probe for Identification of the Take-All Fungus, Gaeumannomyces graminis

    OpenAIRE

    Henson, Joan M.

    1989-01-01

    A 4.3-kilobase mitochondrial DNA fragment was cloned from Gaeumannomyces graminis var. tritici, the causative agent of take-all disease of wheat. Although this DNA fragment hybridized with all three varieties of G. graminis, it showed little homology with DNA from other fungi and thus should be useful for identification of Gaeumannomyces sp. recovered from infected plants.

  4. DNA as a molecular local thermal probe for the analysis of magnetic hyperthermia.

    Science.gov (United States)

    Dias, Jorge T; Moros, María; Del Pino, Pablo; Rivera, Sara; Grazú, Valeria; de la Fuente, Jesus M

    2013-10-25

    Too hot to handle: The surroundings of magnetic nanoparticles can be heated by applying a magnetic field. Polymer-coated magnetic nanoparticles were functionalized with single-stranded DNA molecules and further hybridized with DNA modified with different fluorophores. By correlating the denaturation profiles of the DNA with the local temperature, temperature gradients for the vicinity of the excited nanoparticles were determined.

  5. DNA Probe for Identification of the Take-All Fungus, Gaeumannomyces graminis

    OpenAIRE

    Henson, Joan M.

    1989-01-01

    A 4.3-kilobase mitochondrial DNA fragment was cloned from Gaeumannomyces graminis var. tritici, the causative agent of take-all disease of wheat. Although this DNA fragment hybridized with all three varieties of G. graminis, it showed little homology with DNA from other fungi and thus should be useful for identification of Gaeumannomyces sp. recovered from infected plants.

  6. A Rotational BODIPY Nucleotide: An Environment-Sensitive Fluorescence-Lifetime Probe for DNA Interactions and Applications in Live-Cell Microscopy.

    Science.gov (United States)

    Dziuba, Dmytro; Jurkiewicz, Piotr; Cebecauer, Marek; Hof, Martin; Hocek, Michal

    2016-01-04

    Fluorescent probes for detecting the physical properties of cellular structures have become valuable tools in life sciences. The fluorescence lifetime of molecular rotors can be used to report on variations in local molecular packing or viscosity. We used a nucleoside linked to a meso-substituted BODIPY fluorescent molecular rotor (dC(bdp)) to sense changes in DNA microenvironment both in vitro and in living cells. DNA incorporating dC(bdp) can respond to interactions with DNA-binding proteins and lipids by changes in the fluorescence lifetimes in the range 0.5-2.2 ns. We can directly visualize changes in the local environment of exogenous DNA during transfection of living cells. Relatively long fluorescence lifetimes and extensive contrast for detecting changes in the microenvironment together with good photostability and versatility for DNA synthesis make this probe suitable for analysis of DNA-associated processes, cellular structures, and also DNA-based nanomaterials.

  7. Real-time electrochemical LAMP: a rational comparative study of different DNA intercalating and non-intercalating redox probes.

    Science.gov (United States)

    Martin, Alexandra; Bouffier, Laurent; Grant, Kathryn B; Limoges, Benoît; Marchal, Damien

    2016-06-20

    We present a comparative study of ten redox-active probes for use in real-time electrochemical loop-mediated isothermal amplification (LAMP). Our main objectives were to establish the criteria that need to be fulfilled for minimizing some of the current limitations of the technique and to provide future guidelines in the search for ideal redox reporters. To ensure a reliable comparative study, each redox probe was tested under similar conditions using the same LAMP reaction and the same entirely automatized custom-made real-time electrochemical device (designed for electrochemically monitoring in real-time and in parallel up to 48 LAMP samples). Electrochemical melt curve analyses were recorded immediately at the end of each LAMP reaction. Our results show that there are a number of intercalating and non-intercalating redox compounds suitable for real-time electrochemical LAMP and that the best candidates are those able to intercalate strongly into ds-DNA but not too much to avoid inhibition of the LAMP reaction. The strongest intercalating redox probes were finally shown to provide higher LAMP sensitivity, speed, greater signal amplitude, and cleaner-cut DNA melting curves than the non-intercalating molecules.

  8. Application of DNA fingerprinting with digoxigenated oligonucleotide probe (CAC)5 to analysis of the genetic variation within Taenia taeniaeformis.

    Science.gov (United States)

    Okamoto, M; Ueda, H; Hayashi, M; Oku, Y; Kurosawa, T; Kamiya, M

    1995-04-01

    DNA from T. taeniaeformis digested with the restriction endonuclease was hybridized with digoxigenated oligonucleotide probe (CAC)5. Metacestode and adult showed same clear multibanding patterns, which were characteristic of multilocus DNA fingerprinting. The fingerprinting patterns were quite different from those of the rodent hosts. Genetic variations in 4 laboratory-reared isolates of T. taeniaeformis, including 3 isolates which have been reported to be indistinguishable by infectivity, morphology and protein composition of metacestode, were investigated using this technique. Each of the 4 isolates exhibited isolate-specific fingerprinting patterns and were easily distinguished from one another, thus it was considered that (CAC)5 was a highly resolvable and informative probe for cestodes. However, it was also indicated that (CAC)5 was so sensitive that applying fingerprinting with (CAC)5 to taxonomical or phylogenetic analysis was limited where habitat of the host was restricted to the small area. In comparison to fingerprinting with 32P-labeled (CAC)5, fingerprinting with digoxigenated (CAC)5 represented more and sharper bands. It was considered that a digoxigenated probe was more useful for genetic analysis of cestodes.

  9. Ethidium bromide as a probe of conformational heterogeneity of DNA in chromatin. The role of histone H1.

    Science.gov (United States)

    Lawrence, J J; Daune, M

    1976-07-27

    The accessibility and the tertiary structure of the DNA inside chromatin were studied by using ethidium bromide (EB) as a fluorescent probe. The exclusion model of binding was refined by introductina a parameter alpha (0less than alpha less than 1) which measures the accessibility of the DNA and by taking into account when necessary the existence of two sets of binding sites. We were thus able to fit predicted and experimental isotherms and then to describe completely EB binding to native or partially histone depleted chromatin under various conditions. Itn native chromatin 95% of the DNA (alpha = 0.95) appears to be accessible to EB but two sets of sites are present. The first one corresponds to alpha = 0.13 and is characterized by an affinity constant which is higher by two orders of magnitude than that relative to pure DNA. The second set corresponds to alpha = 0.82 and the corresponding binding constant is only three or four times lower than that of pure DNA. The sites with high affinity are still present after treatment with formaldehyde but disappear after removal of histon H1. By comparison with chromatin treated with deoxycholate of with artifical complexes between H1 and DNA, high affinity sites were found only when all of the histons are bound to DNA. An alpha value around 0.8 is still obtained in 1 M NaC1 treated chromatin, pointing to the fact that histones H3 and H4 are preventing 20% of the DNA to intercalate EB.

  10. Detection of DNA damage by using hairpin molecular beacon probes and graphene oxide.

    Science.gov (United States)

    Zhou, Jie; Lu, Qian; Tong, Ying; Wei, Wei; Liu, Songqin

    2012-09-15

    A hairpin molecular beacon tagged with carboxyfluorescein in combination with graphene oxide as a quencher reagent was used to detect the DNA damage by chemical reagents. The fluorescence of molecular beacon was quenched sharply by graphene oxide; while in the presence of its complementary DNA the quenching efficiency decreased because their hybridization prevented the strong adsorbability of molecular beacon on graphene oxide. If the complementary DNA was damaged by a chemical reagent and could not form intact duplex structure with molecular beacon, more molecular beacon would adsorb on graphene oxide increasing the quenching efficiency. Thus, damaged DNA could be detected based on different quenching efficiencies afforded by damaged and intact complementary DNA. The damage effects of chlorpyrifos-methyl and three metabolites of styrene such as mandelieaeids, phenylglyoxylieaeids and epoxystyrene on DNA were studied as models. The method for detection of DNA damage was reliable, rapid and simple compared to the biological methods. Copyright © 2012 Elsevier B.V. All rights reserved.

  11. Dot-Blot Hybridization for Detection of Five Cucurbit Viruses by Digoxigenin-Labelled cDNA Probes

    Institute of Scientific and Technical Information of China (English)

    MENG Juan; GU Qin-sheng; LIN Shi-ming; PENG Bin; LIU Li-feng; TIAN Yan-ping; LI Li

    2007-01-01

    Dot-blot hybridization was applied in this paper to detect five viruses infecting cucurbitaceous crops,Zuccini yellow mosaic virus(ZYMV),Watermelon mosaic virus(WMV),Cucumber mosaic virus(CMV),Papaya ringspot virus watermelon strain(PRSV-W)and Squash mosaic virus(SqMV),as a good alternative assay in seed health test and epidemiological and transgenic research.Digoxigenin-labelled cDNA probes of the five viruses were synthesized by PCR with the specific primers and applied in dot-blot hybridization to detect five viruses in crude extraction of the infected leaves.And three SqMV probes of different lengths(0.55,1.6,and 2.7 kb,respectively)were designed to investigate the effect of hybridization.The results showed that the sensitivity for detecting the crude extraction of infected leaves by ZYMV,WMV,CMV,PRSV-W,and SqMV was down to 1:160,1:160,1:320,1:160,and 1:320,respectively.Three SqMV probes of different length showed no differences on the sensitivity and specificity.The digoxigenin-labelled probes prepared by PCR could be used for accurate and rapid identification of 5 viruses infecting cucurbitaceous crops with good stabilities,sensitivities,specificity,and reproducibilities.

  12. Methods of staining target chromosomal DNA employing high complexity nucleic acid probes

    Energy Technology Data Exchange (ETDEWEB)

    Gray, Joe W.; Pinkel, Daniel; Kallioniemi, Ol' li-Pekka; Kallioniemi, Anne; Sakamoto, Masaru

    2006-10-03

    Methods and compositions for staining based upon nucleic acid sequence that employ nucleic acid probes are provided. Said methods produce staining patterns that can be tailored for specific cytogenetic analyses. Said probes are appropriate for in situ hybridization and stain both interphase and metaphase chromosomal material with reliable signals. The nucleic acid probes are typically of a complexity greater than 50 kb, the complexity depending upon the cytogenetic application. Methods and reagents are provided for the detection of genetic rearrangements. Probes and test kits are provided for use in detecting genetic rearrangements, particularly for use in tumor cytogenetics, in the detection of disease related loci, specifically cancer, such as chronic myelogenous leukemia (CML), retinoblastoma, ovarian and uterine cancers, and for biological dosimetry. Methods and reagents are described for cytogenetic research, for the differentiation of cytogenetically similar but genetically different diseases, and for many prognostic and diagnostic applications.

  13. Carbon-13 (13C) labeling of Bacillus subtilis vegetative cells and spores: suitability for DNA stable isotope probing (DNA-SIP) of spores in soils.

    Science.gov (United States)

    Nicholson, Wayne L; Fedenko, Jeffrey; Schuerger, Andrew C

    2009-07-01

    To test the suitability of DNA stable isotope probing (DNA-SIP) for characterizing bacterial spore populations in soils, the properties of Bacillus subtilis cells and spores intensely labeled with [(13)C]glucose were characterized. Spore germination, vegetative growth rates, and sporulation efficiency were indistinguishable on glucose versus [(13)C]glucose, as were spore wet heat and UV resistance. Unlabeled and (13)C-labeled spores contained 1.0989 and 74.336 at.% (13)C, and exhibited wet densities of 1.356 and 1.365 g/ml, respectively. Chromosomal DNAs containing (12)C versus (13)C were readily separated by their different buoyant densities in cesium chloride/ethidium bromide gradients.

  14. Isolation and characterization of species-specific DNA probes from Taenia solium and Taenia saginata and their use in an egg detection assay.

    Science.gov (United States)

    Chapman, A; Vallejo, V; Mossie, K G; Ortiz, D; Agabian, N; Flisser, A

    1995-05-01

    Cysticercosis results from ingestion of the eggs of the tapeworm Taenia solium. Reduction of the incidence of human and swine cysticercosis requires identification and treatment of individuals who carry the adult tapeworm. T. solium and Taenia saginata eggs cannot be differentiated on the basis of morphology; thus, in order to improve existing methods for the diagnosis of taeniasis, we have developed highly sensitive, species-specific DNA probes which differentiate T. solium and T. saginata. Recombinant clones containing repetitive DNA sequences which hybridize specifically with genomic DNAs from either species were isolated and characterized. T. solium-specific DNA sequences contained complete and truncated forms of a tandemly repeated 158-bp DNA sequence. An unrelated T. saginata DNA sequence was also characterized and shown to encode a portion of the mitochondrial cytochrome c oxidase I gene. T. solium- and T. saginata-specific DNA probes did not hybridize in dot blot assays either with genomic DNA from the platyhelminths Taenia hydatigena, Taenia pisiformis, Taenia taeniaeformis, Echinococcus granulosus, and Schistosoma mansoni or with genomic DNA from other eukaryotes, including Saccharomyces cerevisiae, Candida albicans, Cryptosporidium parvum, Entamoeba histolytica, Trypanosoma gambiense, Trypanosoma brucei, and Giardia lamblia, Caenorhabditis elegans, and human DNA. By using these T. solium and T. saginata DNA probes, a rapid, highly sensitive and specific dot blot assay for the detection of T. solium eggs was developed.

  15. B to Z-Dna Transition Probed by the Feoli's Formalism for a Kirchhoff Model

    Science.gov (United States)

    Yavari, Morteza

    2013-09-01

    In this paper, we are going to use the techniques of quantum field theory to describe the free energy of DNA structures. The exact solutions of the general equilibrium shape equations for DNA structures in a Kirchhoff model by using the Feoli's formalism [A. Feoli, V. V. Nesterenko and G. Scarpetta, Nucl. Phys. B705, 577 (2005)] are investigated. The free energy of the B- to Z-DNA transition is also calculated in this formalism.

  16. A DNA minor groove electronegative potential genome map based on photo-chemical probing

    DEFF Research Database (Denmark)

    Lindemose, Søren; Nielsen, Peter Eigil; Hansen, Morten

    2011-01-01

    The double-stranded DNA of the genome contains both sequence information directly relating to the protein and RNA coding as well as functional and structural information relating to protein recognition. Only recently is the importance of DNA shape in this recognition process being fully appreciated...... resolution of any genome, and it is illustrated how such detailed studies of this sequence dependent, inherent property of the DNA may reflect on genome organization, gene expression and chromosomal condensation....

  17. Tracking heterotrophic and autotrophic carbon cycling by magnetotactic bacteria in freshwater sediments using DNA stable isotope probing

    Science.gov (United States)

    Kürşat Coşkun, Ömer; Roud, Sophie; He, Kuang; Petersen, Nikolai; Gilder, Stuart; Orsi, William D.

    2017-04-01

    Magnetotactic bacteria (MTB) are diverse, widespread, motile prokaryotes which biomineralize nanosize magnetic minerals, either magnetite or gregite, under highly conserved genetic control and have magnetotaxis to align their position in aquatic environment according to Earth's magnetic field. They play important roles on some geobiological cycle of important minerals such as iron, sulphur, nitrogen and carbon. Yet, to date, their importance in carbon cycle and carbon source in their natural environment have not been previously studied. In this study, we focused on freshwater benthic carbon cycling of MTB and total bacteria using DNA stable isotope probing (DNA-SIP) technique coupled with quantitative PCR (qPCR). Pond sediments from Unterlippach (Germany) were amended with 13C-labelled sodium bicarbonate and 13C-labelled organic matter, and incubated in the dark over a two week time period. Applying separate qPCR assays specific for total bacteria and MTB, respectively, allowed us to estimate the contribution of MTB to total heterotrophic and autotrophic carbon cycling via DNA-SIP. After one week, there was a slight degree of autotrophic activity which increased markedly after two weeks. Comparing total DNA to the qPCR data revealed that changes in the buoyant density of DNA was due mainly to autotrophic bacterial production. DNA-SIP also identified heterotrophic utilization of 13C-labelled organic matter by MTB after 1 week. The qPCR data also allowed us to estimate uptake rates based on the incubation times for heterotrophic and autotrophic MTB. High-throughput DNA sequencing of 16S rRNA genes showed that most of the MTB involved in carbon cycling were related to the Magnetococcus genus. This study sheds light on the carbon sources for MTB in a natural environment and helps unravel their ecological role in the carbon cycle.

  18. Probing Electron-Induced Bond Cleavage at the Single-Molecule Level Using DNA Origami Templates

    DEFF Research Database (Denmark)

    Keller, Adrian Clemens; Bald, Ilko; Rotaru, Alexandru

    2012-01-01

    Low-energy electrons (LEEs) play an important role in nanolithography, atmospheric chemistry, and DNA radiation damage. Previously, the cleavage of specific chemical bonds triggered by LEEs has been demonstrated in a variety of small organic molecules such as halogenated benzenes and DNA nucleoba......Low-energy electrons (LEEs) play an important role in nanolithography, atmospheric chemistry, and DNA radiation damage. Previously, the cleavage of specific chemical bonds triggered by LEEs has been demonstrated in a variety of small organic molecules such as halogenated benzenes and DNA...

  19. Evaluation of the Tissue Culture Standard and Correlation with DNA probes and ELISA for the Detection of Chlamydia Trachomatis

    Science.gov (United States)

    1988-08-01

    reactivity between Chlamydiazyme and Acinetobacter strains. N. Engl. J. Med. 314:922-923. 120. Schachter, J. 1978. Chlamydial infections (first of three...COVEREDL.. TAIW2r)M’ , PtN)Yl COcNIQLTlcIOtU \\JIT OI ’.JP O9606 5 A9J MS THESIS ELIcA F-ot -Tjlt OiiTiio J oi Co- LA ’) OHA T fk v O 4A T S 6...Tissue Culture Standard, and Correlation with DNA Probes and ELISA for the Detection of Chlamydia trachomatis A thesis submitted in partial fulfillment

  20. Identification of PCR-amplified genetically modified organisms (GMOs) DNA by peptide nucleic acid (PNA) probes in anion-exchange chromatographic analysis.

    Science.gov (United States)

    Rossi, Stefano; Lesignoli, Francesca; Germini, Andrea; Faccini, Andrea; Sforza, Stefano; Corradini, Roberto; Marchelli, Rosangela

    2007-04-04

    PCR products obtained by selective amplification of transgenic DNA derived from food samples containing Roundup Ready soybean or Bt-176 maize have been analyzed by anion-exchange HPLC. Peptide nucleic acids (PNAs), oligonucleotide analogues known to bind to complementary single-stranded DNA with high affinity and specificity, have been used as specific probes in order to assess the identity of the peaks observed. Two different protocols were adopted in order to obtain single-stranded DNA: amplification with an excess of one primer or digestion of one DNA strand. The single-stranded DNA was mixed with the PNA probe, and the presence of a specific sequence was revealed through detection of the corresponding PNA:DNA peak with significantly different retention time. Advantages and limits of this approach are discussed. The method was tested with reference materials and subsequently applied to commercial samples.

  1. Detecting variants with Metabolic Design, a new software tool to design probes for explorative functional DNA microarray development

    Directory of Open Access Journals (Sweden)

    Gravelat Fabrice

    2010-09-01

    Full Text Available Abstract Background Microorganisms display vast diversity, and each one has its own set of genes, cell components and metabolic reactions. To assess their huge unexploited metabolic potential in different ecosystems, we need high throughput tools, such as functional microarrays, that allow the simultaneous analysis of thousands of genes. However, most classical functional microarrays use specific probes that monitor only known sequences, and so fail to cover the full microbial gene diversity present in complex environments. We have thus developed an algorithm, implemented in the user-friendly program Metabolic Design, to design efficient explorative probes. Results First we have validated our approach by studying eight enzymes involved in the degradation of polycyclic aromatic hydrocarbons from the model strain Sphingomonas paucimobilis sp. EPA505 using a designed microarray of 8,048 probes. As expected, microarray assays identified the targeted set of genes induced during biodegradation kinetics experiments with various pollutants. We have then confirmed the identity of these new genes by sequencing, and corroborated the quantitative discrimination of our microarray by quantitative real-time PCR. Finally, we have assessed metabolic capacities of microbial communities in soil contaminated with aromatic hydrocarbons. Results show that our probe design (sensitivity and explorative quality can be used to study a complex environment efficiently. Conclusions We successfully use our microarray to detect gene expression encoding enzymes involved in polycyclic aromatic hydrocarbon degradation for the model strain. In addition, DNA microarray experiments performed on soil polluted by organic pollutants without prior sequence assumptions demonstrate high specificity and sensitivity for gene detection. Metabolic Design is thus a powerful, efficient tool that can be used to design explorative probes and monitor metabolic pathways in complex environments

  2. Probing the Conformational Landscape of DNA Polymerases Using Diffusion-Based Single-Molecule FRET

    NARCIS (Netherlands)

    Hohlbein, J.; Kapanidis, A.N.

    2016-01-01

    Monitoring conformational changes in DNA polymerases using single-molecule Förster resonance energy transfer (smFRET) has provided new tools for studying fidelity-related mechanisms that promote the rejection of incorrect nucleotides before DNA synthesis. In addition to the previously known open

  3. Preclinical detection of porcine circovirus type 2 infection using an ultrasensitive nanoparticle DNA probe-based PCR assay.

    Directory of Open Access Journals (Sweden)

    Yong Huang

    Full Text Available Porcine circovirus type 2 (PCV2 has emerged as one of the most important pathogens affecting swine production globally. Preclinical identification of PCV2 is very important for effective prophylaxis of PCV2-associated diseases. In this study, we developed an ultrasensitive nanoparticle DNA probe-based PCR assay (UNDP-PCR for PCV2 detection. Magnetic microparticles coated with PCV2 specific DNA probes were used to enrich PCV2 DNA from samples, then gold nanoparticles coated with PCV2 specific oligonucleotides were added to form a sandwich nucleic acid-complex. After the complex was formed, the oligonucleotides were released and characterized by PCR. This assay exhibited about 500-fold more sensitive than conventional PCR, with a detection limit of 2 copies of purified PCV2 genomic DNA and 10 viral copies of PCV2 in serum. The assay has a wide detection range for all of PCV2 genotypes with reliable reproducibility. No cross-reactivity was observed from the samples of other related viruses including porcine circovirus type 1, porcine parvovirus, porcine pseudorabies virus, porcine reproductive and respiratory syndrome virus and classical swine fever virus. The positive detection rate of PCV2 specific UNDP-PCR in 40 preclinical field samples was 27.5%, which appeared greater than that by conventional and real-time PCR and appeared application potency in evaluation of the viral loads levels of preclinical infection samples. The UNDP-PCR assay reported here can reliably rule out false negative results from antibody-based assays, provide a nucleic acid extraction free, specific, ultrasensitive, economic and rapid diagnosis method for preclinical PCV2 infection in field, which may help prevent large-scale outbreaks.

  4. Spectroscopic probe of the competitive binding of ethidium bromide and neomycin to DNA

    Science.gov (United States)

    Pal, Medini Kanta; Ghosh, Jimut Kanti

    1995-03-01

    The three spectroscopic changes of ethidium bromide (EB) on its binding to DNA, namely red-shift of the νmax, enhancement of fluorescence and induced dichroism are utilized to study the competitive binding of neomycin (NMC) and EB to DNA. Reversion of νmax, decrease in fluorescence and reduction of dichroism of DNA-EB on addition of NMC shows that the binding of NMC and EB to DNA is competitive in nature, over a limited concentration of the polymer. The binding constant of EB-DNA falls from 4.00 × 10 6 to 2.27 × 10 4 1 mol -1 in the presence of added NMC.

  5. Hybridization study of developmental plastid gene expression in mustard (Sinapsis alba L.) with cloned probes for most plastid DNA regions.

    Science.gov (United States)

    Link, G

    1984-07-01

    An approach to assess the extent of developmental gene expression of various regions of plastid (pt)DNA in mustard (Sinapis alba L.) is described. It involves cloning of most ptDNA regions. The cloned regions then serve as hybridization probes to detect and assess the abundance of complementary RNA sequences represented in total plastid RNA. By comparison of the hybridization pattern observed with plastid RNA from either dark-grown or light-grown plants it was found that many ptDNA regions are constitutively expressed, while several 'inducible' regions account for much higher transcript levels in the chloroplast than in the etioplast stage. The reverse situation, i.e. 'repressed' regions which would account for higher transcript levels in the etioplast, was not observed. The hybridization results obtained with RNA from 'intermediatetype' plastids suggest that transient gene expression is a common feature during light-induced chloroplast development. The time-course of gene expression differs for various ptDNA regions.

  6. Application of steered molecular dynamics (SMD) to study DNA-drug complexes and probing helical propensity of amino acids

    Energy Technology Data Exchange (ETDEWEB)

    Orzechowski, Marek [Faculty of Chemistry, Warsaw University, 1 Pasteura Street, Warsaw, 02-093 (Poland); Cieplak, Piotr [Accelrys Incorporated, 9685 Scranton Road, San Diego, CA 92121 (United States)

    2005-05-11

    We present the preliminary results of two computer experiments involving the application of an external force to molecular systems. In the first experiment we simulated the process of pulling out a simple intercalator, the 9-aminoacridine molecule, from its complex with a short DNA oligonucleotide in aqueous solution. Removing a drug from the DNA is assumed to be an opposite process to the complex formation. The force and energy profiles suggest that formation of the DNA-9-aminoacridine complex is preferred when the acridine approaches the DNA from the minor groove rather than the major groove side. For a given mode of pulling the intercalation process is also shown to be nucleotide sequence dependent. In another computer experiment we performed a series of molecular dynamics simulations for stretching short, containing 15 amino acids, helical polypeptides in aqueous solution using an external force. The purpose of these simulations is to check whether this type of approach is sensitive enough to probe the sequence dependent helical propensity of short polypeptides.

  7. Theoretical and Instrumental Studies of the Competitive Interaction Between Aromatic α-Aminobisphosphonates with DNA Using Binding Probes.

    Science.gov (United States)

    Gholivand, M B; Peyman, H; Gholivand, Kh; Roshanfekr, H; Taherpour, A A; Yaghobi, R

    2017-07-01

    Fluorescence spectroscopy, UV-visible absorption spectroscopy, circular dichroism (CD) spectroscopy, viscometry, cyclic voltammetry (CV), and differential pulse voltammetry (DPV) were applied to investigate the competitive interaction of DNA with two aromatic α-aminobisphosphonates and neutral red dye (NR, intercalator) and Hoechst (Ho, groove binder) as spectroscopic probes, in a Tris-hydrogen chloride buffer solution (pH 7.4). The principal component analysis (PCA) was applied to determine the number of chemical components presented in complexation equilibrium of DNA with the aromatic α-aminobisphosphonates (B1 and B2). The spectroscopic and voltammetric studies showed that the groove binding mode of interaction is predominant in the solution containing DNA and α-aminobisphosphonates. Furthermore, the results indicated that α-aminobisphosphonate with the lengthy N-alkyl chains had a stronger interaction. The PCA and theoretical quantum mechanical and molecular mechanic methods were also utilized to determine the structure of DNA with the two α-aminobisphosphonates (B1 and B2).

  8. Mechanism of intramolecular charge transfer in DNA helix as probed by the use of the fluorescent 2-aminopurine

    Institute of Scientific and Technical Information of China (English)

    ZHANG Huijuan; WANG Peng; WANG Xuefei; FENG Juan; XU Sichuan; AI Xicheng; ZHANG Xingkang; ZHANG Jianping

    2004-01-01

    As a structural analogue of adenine, 2-aminopurine (2Ap) is often used as a fluorescent probe to study the intramolecular charge transfer reaction in DNA. We have designed and synthesized a series of model DNA helix with the variation in the distance between the 2Ap probe and the GGG sequence, and have investigated, by means of picosecond time-resolved fluorescence spectroscopy, the effect of the length of the bridge (consisting of a number of transfer dynamics. The fluorescence dynamics of 2Ap exhibited three exponential decay components, the one with a time constant of a few hundred picoseconds is assigned to the intramolecular charge transfer from GGG to 2Ap. Within 2.4 nm of the donor-acceptor separation,the rate of charge transfer decreased exponentially upon increasing the separation, from which the decay factor ,β is determined to be 1.3 nm-1. Beyond 2.4 nm, however, the rate started to increase, this abnormal behavior of charge transfer is interpreted in terms of the match of electronic energies between the I-bridge and the donor/acceptor couple.

  9. DNA flexibility on short length scales probed by atomic force microscopy.

    Science.gov (United States)

    Mazur, Alexey K; Maaloum, Mounir

    2014-02-14

    Unusually high bending flexibility has been recently reported for DNA on short length scales. We use atomic force microscopy (AFM) in solution to obtain a direct estimate of DNA bending statistics for scales down to one helical turn. It appears that DNA behaves as a Gaussian chain and is well described by the wormlike chain model at length scales beyond 3 helical turns (10.5 nm). Below this threshold, the AFM data exhibit growing noise because of experimental limitations. This noise may hide small deviations from the Gaussian behavior, but they can hardly be significant.

  10. Electrochemical DNA Biosensor Based on a Tetrahedral Nanostructure Probe for the Detection of Avian Influenza A (H7N9) Virus.

    Science.gov (United States)

    Dong, Shibiao; Zhao, Rongtao; Zhu, Jiangong; Lu, Xiao; Li, Yang; Qiu, Shaofu; Jia, Leili; Jiao, Xiong; Song, Shiping; Fan, Chunhai; Hao, RongZhang; Song, HongBin

    2015-04-29

    A DNA tetrahedral nanostructure-based electrochemical biosensor was developed to detect avian influenza A (H7N9) virus through recognizing a fragment of the hemagglutinin gene sequence. The DNA tetrahedral probe was immobilized onto a gold electrode surface based on self-assembly between three thiolated nucleotide sequences and a longer nucleotide sequence containing complementary DNA to hybridize with the target single-stranded (ss)DNA. The captured target sequence was hybridized with a biotinylated-ssDNA oligonucleotide as a detection probe, and then avidin-horseradish peroxidase was introduced to produce an amperometric signal through the interaction with 3,3',5,5'-tetramethylbenzidine substrate. The target ssDNA was obtained by asymmetric polymerase chain reaction (PCR) of the cDNA template, reversely transcribed from the viral lysate of influenza A (H7N9) virus in throat swabs. The results showed that this electrochemical biosensor could specifically recognize the target DNA fragment of influenza A (H7N9) virus from other types of influenza viruses, such as influenza A (H1N1) and (H3N2) viruses, and even from single-base mismatches of oligonucleotides. Its detection limit could reach a magnitude of 100 fM for target nucleotide sequences. Moreover, the cycle number of the asymmetric PCR could be reduced below three with the electrochemical biosensor still distinguishing the target sequence from the negative control. To the best of our knowledge, this is the first report of the detection of target DNA from clinical samples using a tetrahedral DNA probe functionalized electrochemical biosensor. It displays that the DNA tetrahedra has a great potential application as a probe of the electrochemical biosensor to detect avian influenza A (H7N9) virus and other pathogens at the gene level, which will potentially aid the prevention and control of the disease caused by such pathogens.

  11. Strain-specific differentiation of lactococci in mixed starter culture populations using randomly amplified polymorphic DNA-derived probes.

    Science.gov (United States)

    Erlandson, K; Batt, C A

    1997-07-01

    A hydrophobic grid membrane filtration (HGMF) colony hybridization assay was developed that allows strain-specific differentiation of defined bacterial populations. The randomly amplified polymorphic DNA (RAPD) fingerprinting technique was used to identify potential signature nucleic acid sequences unique to each member of a commercial cheese starter culture blend. The blend consisted of two closely related Lactococcus lactis subsp. cremoris strains, 160 and 331, and one L. lactis subsp. lactis strain, 210. Three RAPD primers (OPX 1, OPX 12, and OPX 15) generated a total of 32 products from these isolates, 20 of which were potential strain-specific markers. Southern hybridization analyses revealed, that the RAPD-generated signature sequences OPX15-0.95 and a 0.36-kb HaeIII fragment of OPX1-1.0b were specific for strains 331 and 210, respectively, within the context of the test starter culture blend. These strain-specific probes were used in a HGMF colony hybridization assay. Colony lysis, hybridization, and nonradioactive detection parameters were optimized to allow specific differentiation and quantitation of the target strains in the mixed starter culture population. When the 210 and 331 probes were tested at their optimal hybridization temperatures against single cultures, they detected 100% of the target strain CFUs, without cross-reactivity to the other strains. The probes for strains 210 and 331 also successfully detected their targets in blended cultures even with a high background of the other two strains.

  12. 21 CFR 870.1230 - Fiberoptic oximeter catheter.

    Science.gov (United States)

    2010-04-01

    ... 21 Food and Drugs 8 2010-04-01 2010-04-01 false Fiberoptic oximeter catheter. 870.1230 Section 870...) MEDICAL DEVICES CARDIOVASCULAR DEVICES Cardiovascular Diagnostic Devices § 870.1230 Fiberoptic oximeter catheter. (a) Identification. A fiberoptic oximeter catheter is a device used to estimate the...

  13. 21 CFR 876.4530 - Gastroenterology-urology fiberoptic retractor.

    Science.gov (United States)

    2010-04-01

    ... 21 Food and Drugs 8 2010-04-01 2010-04-01 false Gastroenterology-urology fiberoptic retractor. 876... SERVICES (CONTINUED) MEDICAL DEVICES GASTROENTEROLOGY-UROLOGY DEVICES Surgical Devices § 876.4530 Gastroenterology-urology fiberoptic retractor. (a) Identification. A gastroenterology-urology fiberoptic...

  14. Role of the external NH2 linker on the conformation of surface immobilized single strand DNA probes and their SERS detection

    Science.gov (United States)

    He, Lijie; Langlet, Michel; Stambouli, Valerie

    2017-03-01

    The conformation and topological properties of DNA single strand probe molecules attached on solid surfaces are important, notably for the performances of devices such as biosensors. Commonly, the DNA probes are tethered to the surface using external linkers such as NH2. In this study, the role and influence of this amino-linker on the immobilization way and conformation of DNA probes on Ag nanoparticle surface is emphasized using Surface Enhanced Raman Spectroscopy (SERS). We compare the SERS spectra and their reproducibility in the case of two groups of DNA polybase probes which are polyA, polyC, polyT, and polyG. In the first group, the polybases exhibit an external NH2 functional linker while in the second group the polybases are NH2-free. The results show that the reproducibility of SERS spectra is enhanced in the case of the first group. It leads us to propose two models of polybase conformation on Ag surface according to the presence or the absence of the external NH2 linker. In the presence of the NH2 external linker, the latter would act as a major anchoring point. As a result, the polybases are much ordered with a less random orientation than in the case of NH2-free polybases. Consequently, in view of further in situ hybridization for biosensing applications, it is strongly recommended to use NH2 linker functionalized DNA probes.

  15. Methods for the identification of mutations in the human phenylalanine hydroxylase gene using DNA probes

    Energy Technology Data Exchange (ETDEWEB)

    Woo, S.L.C.; Dilella, A.G.

    1990-10-23

    This patent describes a method of detecting a mutation in a phenylalanine hydroxylase gene of human genomic DNA. Also described is an automated method of detecting PKU affected, PKU helerozgotes and normals in fetal to adult human samples.

  16. Probing the Absorption and Emission Transition Dipole Moment of DNA Stabilized Silver Nanoclusters

    DEFF Research Database (Denmark)

    Hooley, Emma Nicole; Carro Temboury, Miguel R.; Vosch, Tom André Jos

    2017-01-01

    Using single molecule polarization measurements, we investigate the excitation and emission polarization characteristics of DNA stabilized silver nanoclusters (C24-AgNCs). Although small changes in the polarization generally accompany changes to the emission spectrum, the emission and excitation ...

  17. DNA as a Model for Probing Polymer Entanglements: Circular Polymers and Non-Classical Dynamics

    Directory of Open Access Journals (Sweden)

    Kathryn Regan

    2016-09-01

    Full Text Available Double-stranded DNA offers a robust platform for investigating fundamental questions regarding the dynamics of entangled polymer solutions. The exceptional monodispersity and multiple naturally occurring topologies of DNA, as well as a wide range of tunable lengths and concentrations that encompass the entanglement regime, enable direct testing of molecular-level entanglement theories and corresponding scaling laws. DNA is also amenable to a wide range of techniques from passive to nonlinear measurements and from single-molecule to bulk macroscopic experiments. Over the past two decades, researchers have developed methods to directly visualize and manipulate single entangled DNA molecules in steady-state and stressed conditions using fluorescence microscopy, particle tracking and optical tweezers. Developments in microfluidics, microrheology and bulk rheology have also enabled characterization of the viscoelastic response of entangled DNA from molecular levels to macroscopic scales and over timescales that span from linear to nonlinear regimes. Experiments using DNA have uniquely elucidated the debated entanglement properties of circular polymers and blends of linear and circular polymers. Experiments have also revealed important lengthscale and timescale dependent entanglement dynamics not predicted by classical tube models, both validating and refuting new proposed extensions and alternatives to tube theory and motivating further theoretical work to describe the rich dynamics exhibited in entangled polymer systems.

  18. DNA mechanics as a tool to probe helicase and translocase activity.

    Science.gov (United States)

    Lionnet, Timothée; Dawid, Alexandre; Bigot, Sarah; Barre, François-Xavier; Saleh, Omar A; Heslot, François; Allemand, Jean-François; Bensimon, David; Croquette, Vincent

    2006-01-01

    Helicases and translocases are proteins that use the energy derived from ATP hydrolysis to move along or pump nucleic acid substrates. Single molecule manipulation has proved to be a powerful tool to investigate the mechanochemistry of these motors. Here we first describe the basic mechanical properties of DNA unraveled by single molecule manipulation techniques. Then we demonstrate how the knowledge of these properties has been used to design single molecule assays to address the enzymatic mechanisms of different translocases. We report on four single molecule manipulation systems addressing the mechanism of different helicases using specifically designed DNA substrates: UvrD enzyme activity detection on a stretched nicked DNA molecule, HCV NS3 helicase unwinding of a RNA hairpin under tension, the observation of RecBCD helicase/nuclease forward and backward motion, and T7 gp4 helicase mediated opening of a synthetic DNA replication fork. We then discuss experiments on two dsDNA translocases: the RuvAB motor studied on its natural substrate, the Holliday junction, and the chromosome-segregation motor FtsK, showing its unusual coupling to DNA supercoiling.

  19. A New FRET-Based Sensitive DNA Sensor for Medical Diagnostics using PNA Probe and Water-Soluble Blue Light Emitting Polymer

    Directory of Open Access Journals (Sweden)

    Nidhi Mathur

    2008-01-01

    Full Text Available A reliable, fast, and low-cost biosensor for medical diagnostics using DNA sequence detection has been developed and tested for the detection of the bacterium “Bacillus anthracis.” In this sensor, Poly [9,9-di (6,6′- N, N′ trimethylammonium hexylfluorenyl-2, 7-diyl-alt-co- (1,4-phenylene] dibromide salt (PFP has been taken as cationic conjugated polymer (CCP and PNA attached with fluorescein dye (PNAC∗ as a probe. The basic principle of this sensor is that when a PNAC∗ probe is hybridized with a single strand DNA (ssDNA having complementary sequence, Forster resonance energy transfer (FRET may take place from PFP to the PNAC∗/DNA complex. If the FRET is efficient, the photoluminescence from the PFP will be highly quenched and that from PNAC∗ will be enhanced. On the other hand, if the DNA sequence is noncomplementary to PNA, FRET will not occur.

  20. U-shaped, double-tapered, fiber-optic sensor for effective biofilm growth monitoring.

    Science.gov (United States)

    Zhong, Nianbing; Zhao, Mingfu; Li, Yishan

    2016-02-01

    To monitor biofilm growth on polydimethylsiloxane in a photobioreactor effectively, the biofilm cells and liquids were separated and measured using a sensor with two U-shaped, double-tapered, fiber-optic probes (Sen. and Ref. probes). The probes' Au-coated hemispherical tips enabled double-pass evanescent field absorption. The Sen. probe sensed the cells and liquids inside the biofilm. The polyimide-silica hybrid-film-coated Ref. probe separated the liquids from the biofilm cells and analyzed the liquid concentration. The biofilm structure and active biomass were also examined to confirm the effectiveness of the measurement using a simulation model. The sensor was found to effectively respond to the biofilm growth in the adsorption through exponential phases at thicknesses of 0-536 μm.

  1. A comparison of DNA damage probes in two HMEC lines withX-irradiation

    Energy Technology Data Exchange (ETDEWEB)

    Wisnewski, Christy L.; Bjornstad, Kathleen A.; Rosen, ChristoperJ.; Chang, Polly Y.; Blakely, Eleanor A.

    2007-01-19

    In this study, we investigated {gamma}H2AX{sup ser139} and 53BP1{sup ser25}, DNA damage pathway markers, to observe responses to radiation insult. Two Human Mammary Epithelial Cell (HMEC) lines were utilized to research the role of immortalization in DNA damage marker expression, HMEC HMT-3522 (S1) with an infinite lifespan, and a subtype of HMEC 184 (184V) with a finite lifespan. Cells were irradiated with 50 cGy X-rays, fixed with 4% paraformaldehyde after 1 hour repair at 37 C, and processed through immunofluorescence. Cells were visualized with a fluorescent microscope and images were digitally captured using Image-Pro Plus software. The 184V irradiated cells exhibited a more positive punctate response within the nucleus for both DNA damage markers compared to the S1 irradiated cells. We will expand the dose and time course in future studies to augment the preliminary data from this research. It is important to understand whether the process of transformation to immortalization compromises the DNA damage sensor and repair process proteins of HMECs in order to understand what is 'normal' and to evaluate the usefulness of cell lines as experimental models.

  2. A COMPARISON OF DNA DAMAGE PROBES IN TWO HMEC LINES WITH X-IRRADIATION

    Energy Technology Data Exchange (ETDEWEB)

    Wisnewski, C.L.; Bjornstad, K.A.; Rosen, C.J.; Chang, P.Y.; Blakely, E.A.

    2007-01-01

    In this study, we investigated γH2AXser139 and 53BP1ser25, DNA damage pathway markers, to observe responses to radiation insult. Two Human Mammary Epithelial Cell (HMEC) lines were utilized to research the role of immortalization in DNA damage marker expression, HMEC HMT-3522 (S1) with an infi nite lifespan, and a subtype of HMEC 184 (184V) with a fi nite lifespan. Cells were irradiated with 50cGy X-rays, fi xed with 4% paraformaldehyde after 1 hour repair at 37°C, and processed through immunofl uorescence. Cells were visualized with a fl uorescent microscope and images were digitally captured using Image-Pro Plus software. The 184V irradiated cells exhibited a more positive punctate response within the nucleus for both DNA damage markers compared to the S1 irradiated cells. The dose and time course will be expanded in future studies to augment the preliminary data from this research. It is important to understand whether the process of transformation to immortalization compromises the DNA damage sensor and repair process proteins of HMECs in order to understand what is “normal” and to evaluate the usefulness of cell lines as experimental models.

  3. Probing the mechanism of recognition of ssDNA by the Cdc13-DBD.

    Science.gov (United States)

    Eldridge, Aimee M; Wuttke, Deborah S

    2008-03-01

    The Saccharomyces cerevisiae protein Cdc13 tightly and specifically binds the conserved G-rich single-stranded overhang at telomeres and plays an essential role in telomere end-protection and length regulation. The 200 residue DNA-binding domain of Cdc13 (Cdc13-DBD) binds an 11mer single-stranded representative of the yeast telomeric sequence [Tel11, d(GTGTGGGTGTG)] with a 3 pM affinity and specificity for three bases (underlined) at the 5' end. The structure of the Cdc13-DBD bound to Tel11 revealed a large, predominantly aromatic protein interface with several unusual features. The DNA adopts an irregular, extended structure, and the binding interface includes a long ( approximately 30 amino acids) structured loop between strands beta2-beta3 (L(2-3)) of an OB-fold. To investigate the mechanism of ssDNA binding, we studied the free and bound states of Cdc13-DBD using NMR spectroscopy. Chemical shift changes indicate that the basic topology of the domain, including L(2-3), is essentially intact in the free state. Changes in slow and intermediate time scale dynamics, however, occur in L(2-3), while conformational changes distant from the DNA interface suggest an induced fit mechanism for binding in the 'hot spot' for binding affinity and specificity. These data point to an overall binding mechanism well adapted to the heterogeneous nature of yeast telomeres.

  4. A Multiplexed, Probe-Based Quantitative PCR Assay for DNA of Phytophthora sojae

    Science.gov (United States)

    Phytophthora sojae (Kaufm. & Gerd.) causes seed rot, pre- and post-emergence damping off, and sometimes foliar blight in soybean (Glycine max). Crop loss may approach 100% with susceptible cultivars. We report here the development of a unique quantitative PCR assay specific to DNA of P. sojae, and a...

  5. Probing the Absorption and Emission Transition Dipole Moment of DNA Stabilized Silver Nanoclusters.

    Science.gov (United States)

    Hooley, Emma N; Carro-Temboury, Miguel R; Vosch, Tom

    2017-02-09

    Using single molecule polarization measurements, we investigate the excitation and emission polarization characteristics of DNA stabilized silver nanoclusters (C24-AgNCs). Although small changes in the polarization generally accompany changes to the emission spectrum, the emission and excitation transition dipoles tend to be steady over time and aligned in a similar direction, when immobilized in PVA. The emission transition dipole patterns, observed for C24-AgNCs in defocused wide field imaging, match that of a single emitter. The small changes to the polarization and spectral shifting that were observed could be due to changes to the conformation of the AgNC or the DNA scaffold. Although less likely, an alternative explanation could be that several well aligned spectrally similar emitters are present within the DNA scaffold which, due to Förster resonance energy transfer (FRET) processes such as energy hopping, energy transfer, and singlet-singlet annihilation, behave as a single emitter. The reported results can provide more insight in the structural and photophysical properties of DNA-stabilized AgNCs.

  6. Chemical grafting of a DNA intercalator probe onto functional iron oxide nanoparticles: a physicochemical study.

    Science.gov (United States)

    Bouffier, Laurent; Yiu, Humphrey H P; Rosseinsky, Matthew J

    2011-05-17

    Spherical magnetite nanoparticles (MNPs, ∼ 24 nm in diameter) were sequentially functionalized with trimethoxysilylpropyldiethylenetriamine (TMSPDT) and a synthetic DNA intercalator, namely, 9-chloro-4H-pyrido[4,3,2-kl]acridin-4-one (PyAcr), in order to promote DNA interaction. The designed synthetic pathway allowed control of the chemical grafting efficiency to access MNPs either partially or fully functionalized with the intercalator moiety. The newly prepared nanomaterials were characterized by a range of physicochemical techniques: FTIR, TEM, PXRD, and TGA. The data were consistent with a full surface coverage by immobilized silylpropyldiethylenetriamine (SPDT) molecules, which corresponds to ∼22,300 SPDT molecules per MNP and a subsequent (4740-2940) PyAcr after the chemical grafting step (i.e., ∼ 2.4 PyAcr/nm(2)). A greater amount of PyAcr (30,600) was immobilized by the alternative strategy of binding a fully prefunctionalized shell to the MNPs with up to 16.1 PyAcr/nm(2). We found that the extent of PyAcr functionalization strongly affects the resulting properties and, particularly, the colloidal stability as well as the surface charge estimated by ζ-potential measurement. The intercalator grafting generates a negative charge contribution which counterbalances the positive charge of the single SPDT shell. The DNA binding capability was measured by titration assay and increases from 15 to 21.5 μg of DNA per mg of MNPs after PyAcr grafting (14-20% yield) but then drops to only ∼2 μg for the fully functionalized MNPs. This highlights that even if the size of the MNPs is obviously a determining factor to promote surface DNA interaction, it is not the only limiting parameter, as the mode of binding and the interfacial charge density are essential to improve loading capability.

  7. Detection of target DNA using fluorescent cationic polymer and peptide nucleic acid probes on solid support

    Directory of Open Access Journals (Sweden)

    Leclerc Mario

    2005-04-01

    Full Text Available Abstract Background Nucleic acids detection using microarrays requires labelling of target nucleic acids with fluorophores or other reporter molecules prior to hybridization. Results Using surface-bound peptide nucleic acids (PNA probes and soluble fluorescent cationic polythiophenes, we show a simple and sensitive electrostatic approach to detect and identify unlabelled target nucleic acid on microarray. Conclusion This simple methodology opens exciting possibilities for applied genetic analysis for the diagnosis of infections, identification of genetic mutations, and forensic inquiries. This electrostatic strategy could also be used with other nucleic acid detection methods such as electrochemistry, silver staining, metallization, quantum dots, or electrochemical dyes.

  8. Development of a 2-Channel Embedded Infrared Fiber-Optic Temperature Sensor Using Silver Halide Optical Fibers

    Directory of Open Access Journals (Sweden)

    Bongsoo Lee

    2011-10-01

    Full Text Available A 2-channel embedded infrared fiber-optic temperature sensor was fabricated using two identical silver halide optical fibers for accurate thermometry without complicated calibration processes. In this study, we measured the output voltages of signal and reference probes according to temperature variation over a temperature range from 25 to 225 °C. To decide the temperature of the water, the difference between the amounts of infrared radiation emitted from the two temperature sensing probes was measured. The response time and the reproducibility of the fiber-optic temperature sensor were also obtained. Thermometry with the proposed sensor is immune to changes if parameters such as offset voltage, ambient temperature, and emissivity of any warm object. In particular, the temperature sensing probe with silver halide optical fibers can withstand a high temperature/pressure and water-chemistry environment. It is expected that the proposed sensor can be further developed to accurately monitor temperature in harsh environments.

  9. The use of radionuclide DNA probe technology for epidemiological studies of tegumentary leishmaniasis in Mato Grosso state

    Directory of Open Access Journals (Sweden)

    Antero Silva Ribeiro de Andrade

    2005-10-01

    Full Text Available DNA hybridisation, using probes labelled with 32P, was used to type Leishmania samples isolated from patients living in endemic areas of Mato Grosso State (Brazil, and clinically diagnosed as having tegumentary leishmaniasis. kDNA cloned mini-circle probes specific for the Leishmania mexicana and Leishmania braziliensis complexes were used. The results showed that L. braziliensis is the predominant group infecting human patients in the state. Sixty-eight samples were typed, 64 samples (94.1% belonging to the L. braziliensis complex and only four (5.9% belonging to the L. mexicana complex. Accurate identification of the Leishmania permits better orientation of the medical follow-up, since clinical manifestations may vary depending on the complex to which the parasite belongs. The epidemiological information furnished by the identification of the Leishmania in given endemic area is also essential for the design of appropriate control measuresHibridização, utilizando sondas de DNA marcadas com 32P, foi utilizada para a tipagem de amostras de Leishmania isoladas de pacientes do estado do Mato Grosso (Brasil, diagnosticados clinicamente como portadores de leishmaniose tegumentar. Sondas de minicírculos clonados de kDNA, específicas para os complexos Leishmania mexicana e Leishmania braziliensis, foram utilizadas. Os resultados demonstraram que o complexo L. brasiliensis é o grupo predominante infectando pacientes humanos no estado do Mato Grosso. Foram tipadas 68 amostras: 64 (94,1% foram identificadas como pertencentes ao complexo L. brasiliensis e somente 4 (5,9% como pertencentes ao complexo L. mexicana. A tipagem de Leishmania é importante para um melhor acompanhamento médico, uma vez que as manifestações clínicas podem variar em função do complexo ao qual o parasita pertence. A informação fornecida pela identificação também é essencial para a definição das medidas de controle mais adequadas e compreensão da epidemiologia da

  10. Identification of DNA sequences from a second pathogenicity island of uropathogenic Escherichia coli CFT073: probes specific for uropathogenic populations.

    Science.gov (United States)

    Rasko, D A; Phillips, J A; Li, X; Mobley, H L

    2001-10-15

    Uropathogenic Escherichia coli is the leading cause of urinary tract infection and hospital visits in North America. Cystitis and acute pyelonephritis, infection of the bladder and kidney, respectively, are the two most common syndromes encountered in patients with urinary tract infection. We sequenced and annotated 71,684 bases of a previously unidentified pathogenicity-associated island (PAI) from E. coli strain CFT073. This PAI contained 89 open-reading frames encoding a pap operon, iron-regulated genes, mobile genetic elements, and a large proportion of unknown or unidentified open-reading frames. Dot blot analysis with 11 DNA sequences from this PAI demonstrated that 7 sequences were more prevalent among uropathogens: 2 probes were more prevalent among cystitis and pyelonephritis isolates, 2 among pyelonephritis isolates only, and 3 among cystitis isolates only than among fecal isolates. These data suggest that groups of uropathogens have genetic differences that may be responsible for the different clinical outcomes.

  11. Standing waves in fiber-optic interferometers

    NARCIS (Netherlands)

    De Haan, V.; Santbergen, R.; Tijssen, M.; Zeman, M.

    2011-01-01

    A study is presented giving the response of three types of fiber-optic interferometers by which a standing wave through an object is investigated. The three types are a Sagnac, Mach–Zehnder and Michelson–Morley interferometer. The response of the Mach–Zehnder interferometer is similar to the Sagnac

  12. Overview of Fiber-Optical Sensors

    Science.gov (United States)

    Depaula, Ramon P.; Moore, Emery L.

    1987-01-01

    Design, development, and sensitivity of sensors using fiber optics reviewed. State-of-the-art and probable future developments of sensors using fiber optics described in report including references to work in field. Serves to update previously published surveys. Systems incorporating fiber-optic sensors used in medical diagnosis, navigation, robotics, sonar, power industry, and industrial controls.

  13. Fluorescent C-linked C8-aryl-guanine probe for distinguishing syn from anti structures in duplex DNA.

    Science.gov (United States)

    Manderville, Richard A; Omumi, Alireza; Rankin née Schlitt, Katherine M; Wilson, Katie A; Millen, Andrea L; Wetmore, Stacey D

    2012-06-18

    The synthesis and optical properties of the carbon (C)-linked C(8)-(2"-benzo[b]thienyl)-2'-deoxyguanosine ((Bth)dG), which acts as a fluorescent reporter of syn versus anti glycosidic conformations in duplex DNA, are described. In the syn-conformation, the probe stabilizes a G:G mismatch, emits at ∼385 nm (excitation ∼285 nm), and shows an induced circular dichroism (ICD) signal at ∼320 nm. Molecular dynamics (MD) simulations predict a wedge (W)-conformation for the mismatched duplex with the C(8)-benzo[b]thienyl moiety residing in the minor groove. In contrast, the probe destabilizes the duplex when base paired with its normal pyrimidine partner C. With flanking purine bases, a major groove B-type duplex is favored with (Bth)dG present in the anti-conformation emitting at ∼413 nm (excitation ∼326 nm) and no ICD signal. However, with flanking pyrimidine bases, (Bth)dG adopts the syn-conformation when base paired with C, and MD simulations predict a base-displaced stacked (S)-conformation, with the opposing C flipped out of the helix. The different duplex (B-, S-, and W-) conformers formed upon incorporation of (Bth)dG are known to play a critical role in the biological activity of N-linked C8-dG adducts formed by arylamine carcinogens. Bulky environment-sensitive fluorescent C(8)-dG adducts that mimic the duplex structures formed by carcinogens may be useful in luminescence-based DNA polymerase assays.

  14. Focused upon hybridization: rapid and high sensitivity detection of DNA using isotachophoresis and peptide nucleic acid probes.

    Science.gov (United States)

    Ostromohov, Nadya; Schwartz, Ortal; Bercovici, Moran

    2015-09-15

    We present a novel assay for rapid and high sensitivity detection of nucleic acids without amplification. Utilizing the neutral backbone of peptide nucleic acids (PNA), our method is based on the design of low electrophoretic mobility PNA probes, which do not focus under isotachophoresis (ITP) unless bound to their target sequence. Thus, background noise associated with free probes is entirely eliminated, significantly improving the signal-to-noise ratio while maintaining a simple single-step assay requiring no amplification steps. We provide a detailed analytical model and experimentally demonstrate the ability to detect targets as short as 17 nucleotides (nt) and a limit of detection of 100 fM with a dynamic range of 5 decades. We also demonstrate that the assay can be successfully implemented for detection of DNA in human serum without loss of signal. The assay requires 15 min to complete, and it could potentially be used in applications where rapid and highly sensitive amplification-free detection of nucleic acids is desired.

  15. PvuII RFLP detected by a human. beta. ADH cDNA probe

    Energy Technology Data Exchange (ETDEWEB)

    Parsian, A.; Burgess, A.K.; Khan, M.A.; Devor, E.J. (Washington Univ. School of Medicine, St. Louis, MO (USA))

    1989-12-11

    A 0.97 kb cDNA (ADH12) fragment encoding human exons 7, 8, 9 of the ADH{sub 2} gene was isolated from an adult human liver cDNA library. The insert can be excised by Pst I digestion. Pvu II identifies a two-allele polymorphism with bands at 4.4 kb (A{sub 1}) and 3.0 kb (A{sub 2}) and invariant bands at 5.1, 4.0, 2.8, and 2.3 kb. It was localized on Chromosome 4q21-q25 by in situ hybridization. Co-dominant segregation was observed in 18 informative families.

  16. Enzyme-Free Detection of Mutations in Cancer DNA Using Synthetic Oligonucleotide Probes and Fluorescence Microscopy

    DEFF Research Database (Denmark)

    Miotke, Laura; Maity, Arindam; Ji, Hanlee

    2015-01-01

    microscopy and nucleic acid analogues have been proposed so far. METHODS AND RESULTS: Here we report a novel enzyme-free approach to efficiently detect cancer mutations. This assay includes gene-specific target enrichment followed by annealing to oligonucleotides containing locked nucleic acids (LNAs...... 1000-fold above the potential detection limit. CONCLUSION: Overall, the novel assay we describe could become a new approach to rapid, reliable and enzyme-free diagnostics of cancer or other associated DNA targets. Importantly, stoichiometry of wild type and mutant targets is conserved in our assay...... of methods since they would allow rapid and relatively inexpensive detection of nucleic acids. Modern fluorescence microscopy is having a huge impact on detection of biomolecules at previously unachievable resolution. However, no straightforward methods to detect DNA in a non-enzymatic way using fluorescence...

  17. Probing the recognition surface of a DNA triplex: Binding studies with intercalator-neomycin conjugates

    OpenAIRE

    XUE Liang; Xi, Hongjuan; Kumar, Sunil; Gray, David; Davis, Erik; Hamilton, Paris; Skirba, Michael; Arya, Dev P.

    2010-01-01

    Thermodynamic studies on the interactions between intercalator-neomycin conjugates and a DNA polynucleotide triplex [poly(dA)•2poly(dT)] were conducted. To draw a complete picture of such interactions, naphthalenedimide-neomycin (3) and anthraquinone-neomycin (4) were synthesized and used together with two other analogues, previously synthesized pyrene-neomycin (1) and BQQ-neomycin (2), in our investigations. A combination of experiments including UV denaturation, circular dichroism (CD) titr...

  18. Characterization of rat brain NCAM mRNA using DNA oligonucleotide probes

    DEFF Research Database (Denmark)

    1990-01-01

    A number of different isoforms of the neural cell adhesion molecule (NCAM) have been identified. The difference between these is due to alternative splicing of a single NCAM gene. In rat brain NCAM mRNAs with sizes of 7.4, 6.7, 5.2, 4.3 and 2.9 kb have been reported. We have synthesized six DNA...... the five NCAM mRNAs in rat brain....

  19. Probing and quantifying DNA-protein interactions with asymmetrical flow field-flow fractionation.

    Science.gov (United States)

    Ashby, Jonathan; Schachermeyer, Samantha; Duan, Yaokai; Jimenez, Luis A; Zhong, Wenwan

    2014-09-05

    Tools capable of measuring binding affinities as well as amenable to downstream sequencing analysis are needed for study of DNA-protein interaction, particularly in discovery of new DNA sequences with affinity to diverse targets. Asymmetrical flow field-flow fractionation (AF4) is an open-channel separation technique that eliminates interference from column packing to the non-covalently bound complex and could potentially be applied for study of macromolecular interaction. The recovery and elution behaviors of the poly(dA)n strand and aptamers in AF4 were investigated. Good recovery of ssDNAs was achieved by judicious selection of the channel membrane with consideration of the membrane pore diameter and the radius of gyration (Rg) of the ssDNA, which was obtained with the aid of a Molecular Dynamics tool. The Rg values were also used to assess the folding situation of aptamers based on their migration times in AF4. The interactions between two ssDNA aptamers and their respective protein components were investigated. Using AF4, near-baseline resolution between the free and protein-bound aptamer fractions could be obtained. With this information, dissociation constants of ∼16nM and ∼57nM were obtained for an IgE aptamer and a streptavidin aptamer, respectively. In addition, free and protein-bound IgE aptamer was extracted from the AF4 eluate and amplified, illustrating the potential of AF4 in screening ssDNAs with high affinity to targets. Our results demonstrate that AF4 is an effective tool holding several advantages over the existing techniques and should be useful for study of diverse macromolecular interaction systems. Copyright © 2014 Elsevier B.V. All rights reserved.

  20. Discovery and validation of DNA hypomethylation biomarkers for liver cancer using HRM-specific probes.

    Directory of Open Access Journals (Sweden)

    Barbara Stefanska

    Full Text Available Poor prognosis of hepatocellular carcinoma (HCC associated with late diagnosis necessitates the development of early diagnostic biomarkers. We have previously delineated the landscape of DNA methylation in HCC patients unraveling the importance of promoter hypomethylation in activation of cancer- and metastasis-driving genes. The purpose of the present study was to test the feasibility that genes that are hypomethylated in HCC could serve as candidate diagnostic markers. We use high resolution melting analysis (HRM as a simple translatable PCR-based method to define methylation states in clinical samples. We tested seven regions selected from the shortlist of genes hypomethylated in HCC and showed that HRM analysis of several of them distinguishes methylation states in liver cancer specimens from normal adjacent liver and chronic hepatitis in the Shanghai area. Such regions were identified within promoters of neuronal membrane glycoprotein M6-B (GPM6B and melanoma antigen family A12 (MAGEA12 genes. Differences in HRM in the immunoglobulin superfamily Fc receptor (FCRL1 separated invasive tumors from less invasive HCC. The identified biomarkers differentiated HCC from chronic hepatitis in another set of samples from Dhaka. Although the main thrust in DNA methylation diagnostics in cancer is on hypermethylated genes, our study for the first time illustrates the potential use of hypomethylated genes as markers for solid tumors. After further validation in a larger cohort, the identified DNA hypomethylated regions can become important candidate biomarkers for liver cancer diagnosis and prognosis, especially in populations with high risk for HCC development.

  1. Coordinate regulation of stromelysin and collagenase genes determined with cDNA probes

    Energy Technology Data Exchange (ETDEWEB)

    Frisch, S.M.; Clark, E.J.; Werb, Z.

    1987-05-01

    Secreted proteinases are required for tumor metastasis, angiogenesis, and tissue remodeling during wound healing and embryonic growth. Thus, the regulation of the genes of secreted proteinases may serve as an interesting model for growth-controlled genes in general. The authors studied the genes of the secreted proteinases stromelysin and collagenase by using molecularly cloned cDNAs from each proteinase. Stromelysin cDNA was cloned by differential screening of a total cDNA library from rabbit synovial cells treated with phorbol 12-myristate 13-acetate, which yielded a clone of 1.2 kilobase pairs; collagenase cDNA was obtained by cloning reverse transcripts of anti-collagenase-immunoadsorbed polysomal mRNA, which yielded a clone of 0.8 kilobase pairs. Stromelysin and collagenase mRNA species of 2.2 and 2.4 kilobases, respectively, were detected on hybridization blots of RNA from phorbol 12-myristate 13-acetate-treated but not untreated rabbit synovial cells. Expression of stromelysin mRNA was also induced in rabbit alveolar macrophages and rabbit brain capillary endothelial cells treated with phorbol 12-myristate 13-acetate. Stromelysin and collagenase mRNA were both induced by phorbol 12-myristate 13-acetate and cytochalasin B at a constant ratio of the two gene products; this suggest coordinate regulation. The fact that induction was blocked after inhibition of protein synthesis by cycloheximide implicates an indirect signal transduction pathway that requires new protein synthesis.

  2. Identification of human DNA in forensic evidence by loop-mediated isothermal amplification combined with a colorimetric gold nanoparticle hybridization probe.

    Science.gov (United States)

    Watthanapanpituck, Khanistha; Kiatpathomchai, Wansika; Chu, Eric; Panvisavas, Nathinee

    2014-11-01

    A DNA test based on loop-mediated isothermal amplification (LAMP) and colorimetric gold nanoparticle (AuNP) hybridization probe to detect the presence of human DNA in forensic evidence was developed. The LAMP primer set targeted eight regions of the human cytochrome b, and its specificity was verified against the DNA of 11 animal species, which included animals closely related to humans, such as chimpanzee and orangutan. By using the AuNP probe, sequence-specific LAMP product could be detected and the test result could be visualized through the change in color. The limit of detection was demonstrated with reproducibility to be as low as 718 fg of genomic DNA, which is equivalent to approximately 100 plasmid DNA copies containing the cytochrome b DNA target region. A simple DNA extraction method for the commonly found forensic biological samples was also devised to streamline the test process. This LAMP-AuNP human DNA test showed to be a robust, specific, and cost-effective tool for the forensic identification of human specimens without requiring sophisticated laboratory instruments.

  3. Identification of metabolically active methanogens in anaerobic digester by DNA Stable-Isotope Probing using 13C-acetate

    Directory of Open Access Journals (Sweden)

    V. Gowdaman

    2015-04-01

    Full Text Available Anaerobic digestion is gaining enormous attention due to the ability to covert organic wastes into biogas, an alternative sustainable energy. Methanogenic community plays a significant role in biogas production and also for proficient functioning of the anaerobic digester. Therefore, this study was carried out to investigate the methanogen diversity of a food waste anaerobic digester. After endogenous respiration, the digester samples were supplemented with isotopes of acetate to enrich methanogen population, and were analyzed using DNA-SIP (Stable-Isotope Probing. Following separation and fractionation of heavy (13C and light (12C DNA, PCR amplification was carried out using archaeal 16S rRNA gene followed by DGGE analysis. Sequencing of the prominent DGGE bands revealed the dominance of Methanocorpusculum labreanum species belonging to hydrogenotrophic Methanomicrobiales, which can produce methane in the presence of H2/CO2 and requires acetate for its growth. This is the first instance where Methanocorpusculum labreanum is being reported as a dominant species in an anaerobic digester operative on food waste.

  4. DNA and protein change in tissues probed by Kubelka-Munk spectral function

    Science.gov (United States)

    Yang, Yuanlong; Celmer, Edward J.; Koutcher, Jason A.; Alfano, Robert R.

    2000-04-01

    Normal, fibroadenoma, malignant, and adipose breast tissues were investigated using Kubelka-Munk Spectral Function (KMSF). The spectral features in KMSF were identified and compared with absorption spectra determined by transmission measurements. A specified spectral feature measured in adipose tissue was assigned to (beta) -carotene, which can be used to separate fat form other molecular components in breast tissues. The peaks of (KMF) at 260nm and 280nm were attributed to DNA and proteins. The signal amplitude over 255nm to 265nm and 275nm to 285nm were found to be different for malignant fibroadenoma, and normal tissues.

  5. GENETIC VARIATION IN RED RASPBERRIES (RUBUS IDAEUS L.; ROSACEAE) FROM SITES DIFFERING IN ORGANIC POLLUTANTS COMPARED WITH SYNTHETIC TANDEM REPEAT DNA PROBES

    Science.gov (United States)

    Two synthetic tandem repetitive DNA probes were used to compare genetic variation at variable-number-tandem-repeat (VNTR) loci among Rubus idaeus L. var. strigosus (Michx.) Maxim. (Rosaceae) individuals sampled at eight sites contaminated by pollutants (N = 39) and eight adjacent...

  6. Using genomic DNA-based probe-selection to improve the sensitivity of high-density oligonucleotide arrays when applied to heterologous species

    Directory of Open Access Journals (Sweden)

    Townsend Henrik J

    2005-11-01

    Full Text Available Abstract High-density oligonucleotide (oligo arrays are a powerful tool for transcript profiling. Arrays based on GeneChip® technology are amongst the most widely used, although GeneChip® arrays are currently available for only a small number of plant and animal species. Thus, we have developed a method to improve the sensitivity of high-density oligonucleotide arrays when applied to heterologous species and tested the method by analysing the transcriptome of Brassica oleracea L., a species for which no GeneChip® array is available, using a GeneChip® array designed for Arabidopsis thaliana (L. Heynh. Genomic DNA from B. oleracea was labelled and hybridised to the ATH1-121501 GeneChip® array. Arabidopsis thaliana probe-pairs that hybridised to the B. oleracea genomic DNA on the basis of the perfect-match (PM probe signal were then selected for subsequent B. oleracea transcriptome analysis using a .cel file parser script to generate probe mask files. The transcriptional response of B. oleracea to a mineral nutrient (phosphorus; P stress was quantified using probe mask files generated for a wide range of gDNA hybridisation intensity thresholds. An example probe mask file generated with a gDNA hybridisation intensity threshold of 400 removed > 68 % of the available PM probes from the analysis but retained >96 % of available A. thaliana probe-sets. Ninety-nine of these genes were then identified as significantly regulated under P stress in B. oleracea, including the homologues of P stress responsive genes in A. thaliana. Increasing the gDNA hybridisation intensity thresholds up to 500 for probe-selection increased the sensitivity of the GeneChip® array to detect regulation of gene expression in B. oleracea under P stress by up to 13-fold. Our open-source software to create probe mask files is freely available http://affymetrix.arabidopsis.info/xspecies/ and may be used to facilitate transcriptomic analyses of a wide range of plant and animal

  7. Probing diversity in freshwater fishes from Mexico and Guatemala with DNA barcodes.

    Science.gov (United States)

    Valdez-Moreno, M; Ivanova, N V; Elías-Gutiérrez, M; Contreras-Balderas, S; Hebert, P D N

    2009-02-01

    The freshwater fish fauna of Mexico and Guatemala is exceptionally diverse with >600 species, many endemic. In this study, patterns of sequence divergence were analysed in representatives of this fauna using cytochrome c oxidase subunit 1 (COI) DNA barcodes for 61 species in 36 genera. The average divergence among conspecific individuals was 0.45%, while congeneric taxa showed 5.1% divergence. Three species of Poblana, each occupying a different crater lake in the arid regions of Central Mexico, have had a controversial taxonomic history but are usually regarded as endemics to a single lake. They possess identical COI barcodes, suggesting a very recent history of isolation. Representatives of the Cichlidae, a complex and poorly understood family, were well discriminated by barcodes. Many species of Characidae seem to be young, with low divergence values (clusters were apparent in the Bramocharax-Astyanax complex. The symbranchid, Opisthernon aenigmaticum, has been regarded as a single species ranging from Guatemala to Mexico, but it includes two deeply divergent barcode lineages, one a possible new endemic species. Aside from these special cases, the results confirm that DNA barcodes will be highly effective in discriminating freshwater fishes from Central America and that a comprehensive analysis will provide new important insights for understanding diversity of this fauna.

  8. Key Structural Elements of Unsymmetrical Cyanine Dyes for Highly Sensitive Fluorescence Turn-On DNA Probes.

    Science.gov (United States)

    Uno, Kakishi; Sasaki, Taeko; Sugimoto, Nagisa; Ito, Hideto; Nishihara, Taishi; Hagihara, Shinya; Higashiyama, Tetsuya; Sasaki, Narie; Sato, Yoshikatsu; Itami, Kenichiro

    2017-01-17

    Unsymmetrical cyanine dyes, such as thiazole orange, are useful for the detection of nucleic acids with fluorescence because they dramatically enhance the fluorescence upon binding to nucleic acids. Herein, we synthesized a series of unsymmetrical cyanine dyes and evaluated their fluorescence properties. A systematic structure-property relationship study has revealed that the dialkylamino group at the 2-position of quinoline in a series of unsymmetrical cyanine dyes plays a critical role in the fluorescence enhancement. Four newly designed unsymmetrical cyanine dyes showed negligible intrinsic fluorescence in the free state and strong fluorescence upon binding to double-stranded DNA (dsDNA) with a quantum yield of 0.53 to 0.90, which is 2 to 3 times higher than previous unsymmetrical cyanine dyes. A detailed analysis of the fluorescence lifetime revealed that the dialkylamino group at the 2-position of quinoline suppressed nonradiative decay in favor of increased fluorescence quantum yield. Moreover, these newly developed dyes were able to stain the nucleus specifically in fixed HeLa cells examined by using a confocal laser-scanning microscope.

  9. Chromatin properties of regulatory DNA probed by manipulation of transcription factors.

    Science.gov (United States)

    Sharov, Alexei A; Nishiyama, Akira; Qian, Yong; Dudekula, Dawood B; Longo, Dan L; Schlessinger, David; Ko, Minoru S H

    2014-08-01

    Transcription factors (TFs) bind to DNA and regulate the transcription of nearby genes. However, only a small fraction of TF binding sites have such regulatory effects. Here we search for the predictors of functional binding sites by carrying out a systematic computational screening of a variety of contextual factors (histone modifications, nuclear lamin-bindings, and cofactor bindings). We used regression analysis to test if contextual factors are associated with upregulation or downregulation of neighboring genes following the induction or knockdown of the 9 TFs in mouse embryonic stem (ES) cells. Functional TF binding sites appeared to be either active (i.e., bound by P300, CHD7, mediator, cohesin, and SWI/SNF) or repressed (i.e., with H3K27me3 histone marks and bound by Polycomb factors). Active binding sites mediated the downregulation of nearby genes upon knocking down the activating TFs or inducing repressors. Repressed TF binding sites mediated the upregulation of nearby genes (e.g., poised developmental regulators) upon inducing TFs. In addition, repressed binding sites mediated repressive effects of TFs, identified by the downregulation of target genes after the induction of TFs or by the upregulation of target genes after the knockdown of TFs. The contextual factors associated with functions of DNA-bound TFs were used to improve the identification of candidate target genes regulated by TFs.

  10. Chromatin Properties of Regulatory DNA Probed by Manipulation of Transcription Factors

    Science.gov (United States)

    Sharov, Alexei A.; Nishiyama, Akira; Qian, Yong; Dudekula, Dawood B.; Longo, Dan L.; Schlessinger, David

    2014-01-01

    Abstract Transcription factors (TFs) bind to DNA and regulate the transcription of nearby genes. However, only a small fraction of TF binding sites have such regulatory effects. Here we search for the predictors of functional binding sites by carrying out a systematic computational screening of a variety of contextual factors (histone modifications, nuclear lamin-bindings, and cofactor bindings). We used regression analysis to test if contextual factors are associated with upregulation or downregulation of neighboring genes following the induction or knockdown of the 9 TFs in mouse embryonic stem (ES) cells. Functional TF binding sites appeared to be either active (i.e., bound by P300, CHD7, mediator, cohesin, and SWI/SNF) or repressed (i.e., with H3K27me3 histone marks and bound by Polycomb factors). Active binding sites mediated the downregulation of nearby genes upon knocking down the activating TFs or inducing repressors. Repressed TF binding sites mediated the upregulation of nearby genes (e.g., poised developmental regulators) upon inducing TFs. In addition, repressed binding sites mediated repressive effects of TFs, identified by the downregulation of target genes after the induction of TFs or by the upregulation of target genes after the knockdown of TFs. The contextual factors associated with functions of DNA-bound TFs were used to improve the identification of candidate target genes regulated by TFs. PMID:24918633

  11. Screening and development of DNA aptamers as capture probes for colorimetric detection of patulin.

    Science.gov (United States)

    Wu, Shijia; Duan, Nuo; Zhang, Weixiao; Zhao, Sen; Wang, Zhouping

    2016-09-01

    Patulin (PAT) is a kind of mycotoxin that has serious harmful impacts on both food quality and human health. A high-affinity ssDNA aptamer that specifically binds to patulin was generated using systemic evolution of ligands by exponential enrichment (SELEX) assisted by graphene oxide (GO). After 15 rounds of positive and negative selection, a highly enriched ssDNA pool was sequenced and the representative sequences were subjected to binding assays to evaluate their affinity and specificity. Of the eight aptamer candidates tested, the sequence PAT-11 bound to patulin with high affinity and excellent selectivity with a dissociation constant (Kd) of 21.83 ± 5.022 nM. The selected aptamer, PAT-11, was subsequently used as a recognition element to develop a detection method for patulin based on an enzyme-chromogenic substrate system. The colorimetric aptasensor exhibited a linear range from 50 to 2500 pg mL(-1), and the limit of detection was found to be 48 pg mL(-1). The results indicated that GO-SELEX technology was appropriate for the screening of aptamers against small-molecule toxins, offering a promising application for aptamer-based biosensors. Copyright © 2016 Elsevier Inc. All rights reserved.

  12. Probing the Influence of Amino Acids on Photoluminescence from Carbon Nanotubes Suspended with DNA.

    Science.gov (United States)

    Kurnosov, N V; Leontiev, V S; Karachevtsev, V A

    2016-11-01

    The quantitative analysis of amino acid levels in the human organism is required for the early clinical diagnosis of a variety of diseases. In this work the influence of 13 amino acid doping on the photoluminescence (PL) from the semiconducting single-walled carbon nanotubes (SWNTs) suspended with single-stranded DNA (ssDNA) in water has been studied. Amino acid doping leads to the PL enhancement and the strongest increase was found after cysteine doping of the nanotube suspension while addition of other amino acids yielded the significantly smaller effect. The emphasis of cysteine molecules is attributed to presence of the reactive thiol group that turns cysteine into reducing agent that passivates the p-defects on the nanotube sidewall and increases the PL intensity. The reasons of PL enhancement after doping with other amino acids are discussed. The response of nanotube PL to cysteine addition depends on the nanotube aqueous suspension preparation with tip or bath sonication treatment. The enhancement of the emission from different nanotube species after cysteine doping was analyzed too. It was shown that the increase of the carbon nanotube PL at addition of cysteine allows successful monitoring of the cysteine concentration in aqueous solution in the range of 50-1000 μM.

  13. Typing for HLA-DPB1*03 and HLA-DPB1*06 using allele-specific DNA in vitro amplification and allele-specific oligonucleotide probes. Detection of "new" DPB1*06 variants

    DEFF Research Database (Denmark)

    Fugger, L; Morling, N; Ryder, L P;

    1989-01-01

    DP gene typing using in vitro DNA amplification combined with sequence-specific oligonucleotide probes has recently been reported. The resulting DNA amplification was specific for the HLA-DPB locus. Typing for the individual DPB alleles was exclusively dependent on the hybridizations of the probe...

  14. U-shaped, double-tapered, fiber-optic sensor for effective biofilm growth monitoring

    Science.gov (United States)

    Zhong, Nianbing; Zhao, Mingfu; Li, Yishan

    2016-01-01

    To monitor biofilm growth on polydimethylsiloxane in a photobioreactor effectively, the biofilm cells and liquids were separated and measured using a sensor with two U-shaped, double-tapered, fiber-optic probes (Sen. and Ref. probes). The probes’ Au-coated hemispherical tips enabled double-pass evanescent field absorption. The Sen. probe sensed the cells and liquids inside the biofilm. The polyimide–silica hybrid-film-coated Ref. probe separated the liquids from the biofilm cells and analyzed the liquid concentration. The biofilm structure and active biomass were also examined to confirm the effectiveness of the measurement using a simulation model. The sensor was found to effectively respond to the biofilm growth in the adsorption through exponential phases at thicknesses of 0–536 μm. PMID:26977344

  15. Phylogenetic relationships of annual and perennial wild rice: probing by direct DNA sequencing.

    Science.gov (United States)

    Barbier, P; Morishima, H; Ishihama, A

    1991-05-01

    The phylogenetic relationships between Asian wild rice strains were analyzed by direct sequencing of PCR-amplified DNA fragments. The sequence of three introns located in the phytochrome gene was determined for eight strains of the Asian wild rice, Oryza rufipogon, and one strain of the related African species, Oryza longistaminata. The number of nucleotide substitutions per site between various strains within a single species, O. rufipogon, ranged between 0.0017 and 0.0050, while those between two related species, O. rufipogon and O. longistaminate, were 0.043-0.049 (23-26 within 532 bp). Taken together with the sequence differences of the 10-kDa prolamin gene, a model is proposed for the phylogenetic relationships and evolutionary history of annuals and perennials within O. rufipogon.

  16. Differentiation of Moraxella nonliquefaciens, M. lacunata, and M. bovis by using multilocus enzyme electrophoresis and hybridization with pilin-specific DNA probes.

    Science.gov (United States)

    Tønjum, T; Caugant, D A; Bøvre, K

    1992-12-01

    Genetic relationships among strains of Moraxella nonliquefaciens, M. lacunata, and M. bovis were studied by using multilocus enzyme electrophoresis and DNA-DNA hybridization. The 74 isolates analyzed for electrophoretic variation at 12 enzyme loci were assigned to 59 multilocus genotypes. The multilocus genotypes were grouped in four major clusters, one representing strains of M. nonliquefaciens, two representing strains of M. lacunata, and one comprising strains of M. bovis and the single strain of M. equi analyzed. DNA-DNA hybridization with total genomic probes also revealed four major distinctive entities that corresponded to those identified by multilocus enzyme electrophoresis. The two distinct clusters recognized among the M. lacunata strains apparently corresponded to the species previously designated M. lacunata and M. liquefaciens. Distinction of the four entities was improved by hybridization with polymerase chain reaction products of nonconserved parts of pilin genes as DNA probes. With these polymerase chain reaction probes, new isolates of M. nonliquefaciens, M. lacunata, M. liquefaciens, and M. bovis can be identified easily by hybridization.

  17. Fiber-Optic Surface Temperature Sensor Based on Modal Interference

    Directory of Open Access Journals (Sweden)

    Frédéric Musin

    2016-07-01

    Full Text Available Spatially-integrated surface temperature sensing is highly useful when it comes to controlling processes, detecting hazardous conditions or monitoring the health and safety of equipment and people. Fiber-optic sensing based on modal interference has shown great sensitivity to temperature variation, by means of cost-effective image-processing of few-mode interference patterns. New developments in the field of sensor configuration, as described in this paper, include an innovative cooling and heating phase discrimination functionality and more precise measurements, based entirely on the image processing of interference patterns. The proposed technique was applied to the measurement of the integrated surface temperature of a hollow cylinder and compared with a conventional measurement system, consisting of an infrared camera and precision temperature probe. As a result, the optical technique is in line with the reference system. Compared with conventional surface temperature probes, the optical technique has the following advantages: low heat capacity temperature measurement errors, easier spatial deployment, and replacement of multiple angle infrared camera shooting and the continuous monitoring of surfaces that are not visually accessible.

  18. Partially reduced graphene oxide based FRET on fiber-optic interferometer for biochemical detection

    Science.gov (United States)

    Yao, B. C.; Wu, Y.; Yu, C. B.; He, J. R.; Rao, Y. J.; Gong, Y.; Fu, F.; Chen, Y. F.; Li, Y. R.

    2016-03-01

    Fluorescent resonance energy transfer (FRET) with naturally exceptional selectivity is a powerful technique and widely used in chemical and biomedical analysis. However, it is still challenging for conventional FRET to perform as a high sensitivity compact sensor. Here we propose a novel ‘FRET on Fiber’ concept, in which a partially reduced graphene oxide (prGO) film is deposited on a fiber-optic modal interferometer, acting as both the fluorescent quencher for the FRET and the sensitive cladding for optical phase measurement due to refractive index changes in biochemical detection. The target analytes induced fluorescence recovery with good selectivity and optical phase shift with high sensitivity are measured simultaneously. The functionalized prGO film coated on the fiber-optic interferometer shows high sensitivities for the detections of metal ion, dopamine and single-stranded DNA (ssDNA), with detection limits of 1.2 nM, 1.3 μM and 1 pM, respectively. Such a prGO based ‘FRET on fiber’ configuration, bridging the FRET and the fiber-optic sensing technology, may serve as a platform for the realization of series of integrated ‘FRET on Fiber’ sensors for on-line environmental, chemical, and biomedical detection, with excellent compactness, high sensitivity, good selectivity and fast response

  19. Standing waves in fiber-optic interferometers.

    Science.gov (United States)

    de Haan, V; Santbergen, R; Tijssen, M; Zeman, M

    2011-10-10

    A study is presented giving the response of three types of fiber-optic interferometers by which a standing wave through an object is investigated. The three types are a Sagnac, Mach-Zehnder and Michelson-Morley interferometer. The response of the Mach-Zehnder interferometer is similar to the Sagnac interferometer. However, the Sagnac interferometer is much harder to study because of the fact that one input port and output port coincide. Further, the Mach-Zehnder interferometer has the advantage that the output ports are symmetric, reducing the systematic effects. Examples of standing wave light absorption in several simple objects are given. Attention is drawn to the influence of standing waves in fiber-optic interferometers with weak-absorbing layers incorporated. A method is described for how these can be theoretically analyzed and experimentally measured. Further experiments are needed for a thorough comparison between theory and experiment.

  20. Fiberoptic metal detector capable of profile detection

    OpenAIRE

    Hua, Wei-Shu; Hooks, Joshua R.; Erwin, Nicholas A.; Wu, Wen-Jong; Wang, Wei-Chih

    2011-01-01

    The purpose of this paper is to develop a novel ferromagnetic polymeric metal detector system by using a fiber-optic Mach-Zehnder interferometer with a newly developed ferromagnetic polymer as the magnetostrictive sensing device. This ferromagnetic polymeric metal detector system is simple to fabricate, small in size, and resistant to RF interference (which is common in typical electromagnetic type metal detectors). Metal detection is made possible by disrupting the magnetic flux density pres...

  1. Probing the recognition surface of a DNA triplex: binding studies with intercalator-neomycin conjugates.

    Science.gov (United States)

    Xue, Liang; Xi, Hongjuan; Kumar, Sunil; Gray, David; Davis, Erik; Hamilton, Paris; Skriba, Michael; Arya, Dev P

    2010-07-06

    Thermodynamic studies on the interactions between intercalator-neomycin conjugates and a DNA polynucleotide triplex [poly(dA).2poly(dT)] were conducted. To draw a complete picture of such interactions, naphthalene diimide-neomycin (3) and anthraquinone-neomycin (4) conjugates were synthesized and used together with two other analogues, previously synthesized pyrene-neomycin (1) and BQQ-neomycin (2) conjugates, in our investigations. A combination of experiments, including UV denaturation, circular dichroism (CD) titration, differential scanning calorimetry (DSC), and isothermal titration calorimetry (ITC), revealed that all four conjugates (1-4) stabilized poly(dA).2poly(dT) much more than its parent compound, neomycin. UV melting experiments clearly showed that the temperature (T(m3-->2)) at which poly(dA).2poly(dT) dissociated into poly(dA).poly(dT) and poly(dT) increased dramatically (>12 degrees C) in the presence of intercalator-neomycin conjugates (1-4) even at a very low concentration (2 muM). In contrast to intercalator-neomycin conjugates, the increment of T(m3-->2) of poly(dA).2poly(dT) induced by neomycin was negligible under the same conditions. The binding preference of intercalator-neomycin conjugates (1-4) to poly(dA).2poly(dT) was also confirmed by competition dialysis and a fluorescent intercalator displacement assay. Circular dichroism titration studies revealed that compounds 1-4 had slightly larger binding site size ( approximately 7-7.5) with poly(dA).2poly(dT) as compared to neomycin ( approximately 6.5). The thermodynamic parameters of these intercalator-neomycin conjugates with poly(dA).2poly(dT) were derived from an integrated van't Hoff equation using the T(m3-->2) values, the binding site size numbers, and other parameters obtained from DSC and ITC. The binding affinity of all tested ligands with poly(dA).2poly(dT) increased in the following order: neomycin intercalator-neomycin conjugates for poly(dA).2poly(dT) increases as a function of

  2. Positively charged polymer brush-functionalized filter paper for DNA sequence determination following Dot blot hybridization employing a pyrrolidinyl peptide nucleic acid probe.

    Science.gov (United States)

    Laopa, Praethong S; Vilaivan, Tirayut; Hoven, Voravee P

    2013-01-07

    As inspired by the Dot blot analysis, a well known technique in molecular biology and genetics for detecting biomolecules, a new paper-based platform for colorimetric detection of specific DNA sequences employing peptide nucleic acid (PNA) as a probe has been developed. In this particular study, a pyrrolidinyl PNA bearing a conformationally rigid d-prolyl-2-aminocyclopentanecarboxylic acid backbone (acpcPNA) was used as a probe. The filter paper was modified to be positively charged with grafted polymer brushes of quaternized poly(dimethylamino)ethyl methacrylate (QPDMAEMA) prepared by surface-initiated polymerization of 2-(dimethylamino)ethyl methacrylate from the filter paper via ARGET ATRP followed by quaternization with methyl iodide. Following the Dot blot format, a DNA target was first immobilized via electrostatic interactions between the positive charges of the QPDMAEMA brushes and negative charges of the phosphate backbone of DNA. Upon hybridization with the biotinylated pyrrolidinyl peptide nucleic acid (b-PNA) probe, the immobilized DNA can be detected by naked eye observation of the yellow product generated by the enzymatic reaction employing HRP-labeled streptavidin. It has been demonstrated that this newly developed assay was capable of discriminating between complementary and single base mismatch targets at a detection limit of at least 10 fmol. In addition, the QPDMAEMA-grafted filter paper exhibited a superior performance to the commercial membranes, namely Nylon 66 and nitrocellulose.

  3. Cultivation-Independent Detection of Autotrophic Hydrogen-Oxidizing Bacteria by DNA Stable-Isotope Probing

    Science.gov (United States)

    Pumphrey, Graham M.; Ranchou-Peyruse, Anthony; Spain, Jim C.

    2011-01-01

    Knallgas bacteria are a physiologically defined group that is primarily studied using cultivation-dependent techniques. Given that current cultivation techniques fail to grow most bacteria, cultivation-independent techniques that selectively detect and identify knallgas bacteria will improve our ability to study their diversity and distribution. We used stable-isotope probing (SIP) to identify knallgas bacteria in rhizosphere soil of legumes and in a microbial mat from Obsidian Pool in Yellowstone National Park. When samples were incubated in the dark, incorporation of 13CO2 was H2 dependent. SIP enabled the detection of knallgas bacteria that were not detected by cultivation, and the majority of bacteria identified in the rhizosphere soils were betaproteobacteria predominantly related to genera previously known to oxidize hydrogen. Bacteria in soil grew on hydrogen at concentrations as low as 100 ppm. A hydB homolog encoding a putative high-affinity NiFe hydrogenase was amplified from 13C-labeled DNA from both vetch and clover rhizosphere soil. The results indicate that knallgas bacteria can be detected by SIP and populations that respond to different H2 concentrations can be distinguished. The methods described here should be applicable to a variety of ecosystems and will enable the discovery of additional knallgas bacteria that are resistant to cultivation. PMID:21622787

  4. 用DNA探针检测沙眼衣原体%Detection of Chlamydia Trachomatis by DNA Probe Blot

    Institute of Scientific and Technical Information of China (English)

    王卫萍; 陈亚利; 武建国

    2001-01-01

    目的: 建立一种敏感而特异的沙眼衣原体分子生物学检测方法. 方法: 用PCR扩增517bp的沙眼衣原体种特异性基因片段并标记成探针,建立DNA探针杂交检测沙眼衣原体的方法.结果: 探针只与沙眼衣原体L2、TE55株DNA呈阳性杂交斑点,与其他两种衣原体、解脲支原体、淋病奈瑟菌、大肠埃希菌、金黄色葡萄球菌、流感嗜血杆菌及白色念珠菌DNA斑点膜无阳性杂交信号.从100例慢性宫颈炎和前列腺炎病人生殖道分泌物中检出阳性22例,阳性率22%. 结论: 建立的DNA探针检测沙眼衣原体方法具有较高的敏感性和特异性,可用于批量临床标本的检测.%Objectives: To establish a sensitive and specific molecular biologicalmethod for detecting Chlamydia trachomatis of genital tract infection patients. Methods: A DNA blot assay was developed by coating DNA of Chlamydia trachomatisand/or extract of clinical samples on nitrocellulose (NC) membrane, blotting witha DNA probe labeled with DIG. Results: There was no positive blotting in Chlamydiapneumoniae, Chlamydia psittaci, Ureaplasma urealyticum, Neisseria gonorrhoeae,Esherichia coli, Staphylococcus aureus, Haemophilus infuenzae and Candida albicansexcept Chlamydia trachomatis. The sensitivity could be improved to 1pg. The positivepercentage was 22% (22/100) in detection of swabs collected from 100 chroniccervicitis and prostatitis patients. Conclusions: This method was not only sensitive, rapid and specific but also could be applied to detect batch samples. Natl J Androl,2001,7(2):102~104

  5. Spectroscopic and molecular modeling methods to investigate the interaction between 5-Hydroxymethyl-2-furfural and calf thymus DNA using ethidium bromide as a probe.

    Science.gov (United States)

    Zhu, Jinhua; Chen, Lanlan; Dong, Yingying; Li, Jiazhong; Liu, Xiuhua

    2014-04-24

    In this work, the interaction of 5-Hydroxymethyl-2-furfural (5-HMF) with calf thymus DNA (ctDNA) under simulated physiological conditions (Tris-HCl buffer of pH 7.40), was explored by UV absorption spectroscopy, fluorescence spectroscopy and molecular modeling method, using ethidium bromide (EB) as a fluorescence probe of DNA. The fluorescence quenching mechanism of EB-ctDNA by 5-HMF was confirmed to be a static quenching, which derived from the formation of a new complex. The binding constants of 5-HMF with DNA in the presence of EB were calculated to be 2.17×10(3), 4.24×10(3) and 6.95×10(3) L mol(-1) at 300, 305 and 310 K, respectively. The calculated thermodynamic parameters, enthalpy change ΔH and entropy change ΔS, suggested that both hydrophobic interactions and hydrogen bonds played a predominant role in the binding of 5-HMF to DNA. According to the UV absorption spectroscopy and melting temperature (Tm) curve results, the binding mode of 5-HMF with DNA was indicative of a non-intercalative binding, which was supposed to be a groove binding. The molecular modeling results showed that 5-HMF could bind into the hydrophobic region of ctDNA and supported the conclusions obtained from the above experiments.

  6. In situ fiber-optical monitoring of cytosolic calcium in tissue explant cultures

    CERN Document Server

    Ryser, Manuel; Geiser, Marianne; Frenz, Martin; Rička, Jaro

    2014-01-01

    We present a fluorescence-lifetime based method for monitoring cell and tissue activity in situ, during cell culturing and in the presence of a strong autofluorescence background. The miniature fiber-optic probes are easily incorporated in the tight space of a cell culture chamber or in an endoscope. As a first application we monitored the cytosolic calcium levels in porcine tracheal explant cultures using the Calcium Green-5N (CG5N) indicator. Despite the simplicity of the optical setup we are able to detect changes of calcium concentration as small as 2.5 nM, with a monitoring time resolution of less than 1 s.

  7. Fiber-optic raster scanning two-photon endomicroscope using a tubular piezoelectric actuator

    Science.gov (United States)

    Do, Dukho; Yoo, Hongki; Gweon, Dae-Gab

    2014-06-01

    A nonresonant, fiber-optic raster scanning endomicroscope was developed using a quarter-tubular piezoelectric (PZT) actuator. A fiber lever mechanism was utilized to enhance the small actuation range of the tubular PZT actuator and to increase its field-of-view. Finite element method simulation of the endoscopic probe was conducted for various conditions to maximize its scanning range. After fabricating the probe using a double clad fiber, we obtained two-photon fluorescence images using raster beam scanning of the fiber. The outer diameter of the probe was 3.5 mm and its rigid distal length was 30 mm including a high numerical aperture gradient index lens. These features are sufficient for input into the instrumental channel of a commercial colonoscope or gastroscope to obtain high resolution images in vivo.

  8. Probing the Interaction between a DNA Nucleotide (Adenosine-5'-Monophosphate Disodium) and Surface Active Ionic Liquids by Rotational Relaxation Measurement and Fluorescence Correlation Spectroscopy.

    Science.gov (United States)

    Roy, Arpita; Banerjee, Pavel; Dutta, Rupam; Kundu, Sangita; Sarkar, Nilmoni

    2016-10-02

    This article demonstrates the interaction of a deoxyribonucleic acid (DNA) nucleotide, adenosine-5'-monophosphate disodium (AMP) with a cationic surface active ionic liquid (SAIL) 1-dodecyl-3-methylimidazoium chloride (C12mimCl) and an anionic SAIL, 1-butyl-3-methylimidazolium n-octylsulfate ([C4mim][C8SO4]). Dynamic light scattering (DLS) measurements and 1H NMR (nuclear magnetic resonance) studies indicate that substantial interaction is taking place among the DNA nucleotide, AMP and the SAILs. Moreover, cryogenic transmission electron microscopy (cryo-TEM) suggests that SAILs containing micellar assemblies are transformed into larger micellar assemblies in presence of DNA nucleotide. Additionally, the rotational motion of two oppositely charged molecules, Rhodamine 6G perchlorate (R6G) and Fluorescein sodium salt (Fl-Na) have been monitored in these aggregates. The rotational motion of R6G and Fl-Na differs significantly between SAILs micelles, and SAILs-AMP containing larger micellar aggregates. The effect of negatively charged DNA nucleotide (AMP) addition into the cationic and anionic SAILs is more prominent for the cationic charged molecule R6G than that of anionic probe Fl-Na due to the favourable electrostatic interaction between the AMP and cationic R6G. Moreover, the influence of the anionic DNA nucleotide on the cationic and anionic SAIL micelles is monitored through the variation of the lateral diffusion motion of oppositely charged probe molecules (R6G and Fl-Na) inside these aggregates. This variation in diffusion coefficient values also suggests that interaction pattern of these oppositely charged probes are different within the SAILs-nucleotide containing aggregates. Therefore, both rotational and translational diffusion measurements confirm that the DNA nucleotide (AMP) renders more rigid microenvironment within the micellar solution of SAILs.

  9. Development of an on-site rapid real-time polymerase chain reaction system and the characterization of suitable DNA polymerases for TaqMan probe technology.

    Science.gov (United States)

    Furutani, Shunsuke; Naruishi, Nahoko; Hagihara, Yoshihisa; Nagai, Hidenori

    2016-08-01

    On-site quantitative analyses of microorganisms (including viruses) by the polymerase chain reaction (PCR) system are significantly influencing medical and biological research. We have developed a remarkably rapid and portable real-time PCR system that is based on microfluidic approaches. Real-time PCR using TaqMan probes consists of a complex reaction. Therefore, in a rapid real-time PCR, the optimum DNA polymerase must be estimated by using actual real-time PCR conditions. In this study, we compared the performance of three DNA polymerases in actual PCR conditions using our rapid real-time PCR system. Although KAPA2G Fast HS DNA Polymerase has the highest enzymatic activity among them, SpeedSTAR HS DNA Polymerase exhibited better performance to rapidly increase the fluorescence signal in an actual real-time PCR using TaqMan probes. Furthermore, we achieved rapid detection of Escherichia coli in 7 min by using SpeedSTAR HS DNA Polymerase with the same sensitivity as that of a conventional thermal cycler.

  10. DNA Detection of Toxoplasma gondii with a Magnetic Molecular Beacon Probe via CdTe@Ni Quantum Dots as Energy Donor

    Directory of Open Access Journals (Sweden)

    Shichao Xu

    2013-01-01

    Full Text Available A new method for detection of Toxoplasma gondii via DNA sensing technology was developed in this study. It was based on the mechanism of fluorescence resonance energy transfer (FRET in which multifunctional and magnetic-fluorescent CdTe@Ni quantum dots (mQDs were utilized as energy donor and a commercial BHQ2 as acceptor. The sensing probe was fabricated by labeling a stem-loop Toxoplasma gondii DNA oligonucleotide with CdTe@Ni mQDs at the 5′ end and BHQ2 at 3′ end, respectively. The surface assembly of CdTe on Ni core and the formation of CdTe@Ni were confirmed by XRD analysis. The sizes of CdTe, Ni nanoparticles, and CdTe@Ni were measured via TEM and XRD methods and estimated to be ~3 nm, ~15 nm, and ~20 nm, respectively. The sensing ability was investigated by the fluorescence spectrum (FS. An obvious fluorescence recovery was observed when the complete complimentary target Toxoplasma gondii DNA was introduced, which did not happen in the case of the target DNA with one-base pair mismatch. Our research indicates that the current sensing probe is sensitive and specific in detection of Toxoplasma gondii DNA and has great potential in Toxoplasmosis diagnosis.

  11. Detecting 22q11.2 deletions by use of multiplex ligation-dependent probe amplification on DNA from neonatal dried blood spot samples

    DEFF Research Database (Denmark)

    Sørensen, Karina M; Agergaard, Peter; Olesen, Charlotte;

    2010-01-01

    of 22q11.2 deletions among certain manifestations, eg, congenital heart disease, on selected Danes, a multiplex ligation-dependant probe amplification (MLPA) analysis was designed. The analysis was planned to be performed on DNA extracted from dried blood spot samples (DBSS) obtained from Guthrie cards...... MLPA design using nine patients diagnosed with the 22q11.2 deletion and 101 controls. All deletions were identified using DNA extracted from DBSS, and no copy number variations were detected in the controls, resulting in a specificity and sensitivity of 100%. It is thereby concluded that the novel MLPA...

  12. Fiber-optic Michelson interferometer using an optical power divider.

    Science.gov (United States)

    Imai, M; Ohashi, T; Ohtsuka, Y

    1980-10-01

    A fiber-optic interferometer consisting of a multimode fiber-optical power divider was constructed in the Michelson arrangement and applied to measure a micrometer-order displacement of the vibrating object based on an optical homodyne technique. Improvement in the sensitivity of the apparatus is discussed from the viewpoint of increasing the minimum detectable beat signal.

  13. Fiber-optic interferometric acoustic sensors for wind tunnel applications

    Science.gov (United States)

    Cho, Y. C.

    1993-01-01

    Progress in developing fiber-optic interferometric sensors for aeroacoustic measurements in wind tunnels, performed under the NASA program, is reported. Preliminary results show that the fiber-optic interferometer sensor array is a powerful instrument for solving complex acoustic measurement problems in wind tunnels, which cannot be resolved with the conventional transducer technique.

  14. Fiber-optic interferometric acoustic sensors for wind tunnel applications

    Science.gov (United States)

    Cho, Y. C.

    1993-01-01

    Progress in developing fiber-optic interferometric sensors for aeroacoustic measurements in wind tunnels, performed under the NASA program, is reported. Preliminary results show that the fiber-optic interferometer sensor array is a powerful instrument for solving complex acoustic measurement problems in wind tunnels, which cannot be resolved with the conventional transducer technique.

  15. Locking-to-unlocking system is an efficient strategy to design DNA/silver nanoclusters (AgNCs) probe for human miRNAs

    DEFF Research Database (Denmark)

    Shah, Pratik; Choi, Suk Won; Kim, Ho-jin;

    2016-01-01

    MicroRNAs (miRNAs), small non-coding RNA molecules, are important biomarkers for research and medical purposes. Here, we describe the development of a fast and simple method using highly fluorescent oligonucleotide-silver nanocluster probes (DNA/AgNCs) to efficiently detect specific miRNAs. Due t...... detect a number of cancer related miRNAs in RNA extracts from human cancer cell lines....

  16. UniPROBE, update 2015: new tools and content for the online database of protein-binding microarray data on protein–DNA interactions

    Science.gov (United States)

    Hume, Maxwell A.; Barrera, Luis A.; Gisselbrecht, Stephen S.; Bulyk, Martha L.

    2015-01-01

    The Universal PBM Resource for Oligonucleotide Binding Evaluation (UniPROBE) serves as a convenient source of information on published data generated using universal protein-binding microarray (PBM) technology, which provides in vitro data about the relative DNA-binding preferences of transcription factors for all possible sequence variants of a length k (‘k-mers’). The database displays important information about the proteins and displays their DNA-binding specificity data in terms of k-mers, position weight matrices and graphical sequence logos. This update to the database documents the growth of UniPROBE since the last update 4 years ago, and introduces a variety of new features and tools, including a new streamlined pipeline that facilitates data deposition by universal PBM data generators in the research community, a tool that generates putative nonbinding (i.e. negative control) DNA sequences for one or more proteins and novel motifs obtained by analyzing the PBM data using the BEEML-PBM algorithm for motif inference. The UniPROBE database is available at http://uniprobe.org. PMID:25378322

  17. Conformation-sensitive nucleoside analogues as topology-specific fluorescence turn-on probes for DNA and RNA G-quadruplexes

    Science.gov (United States)

    Tanpure, Arun A.; Srivatsan, Seergazhi G.

    2015-01-01

    Development of probes that can discriminate G-quadruplex (GQ) structures and indentify efficient GQ binders on the basis of topology and nucleic acid type is highly desired to advance GQ-directed therapeutic strategies. In this context, we describe the development of minimally perturbing and environment-sensitive pyrimidine nucleoside analogues, based on a 5-(benzofuran-2-yl)uracil core, as topology-specific fluorescence turn-on probes for human telomeric DNA and RNA GQs. The pyrimidine residues of one of the loop regions (TTA) of telomeric DNA and RNA GQ oligonucleotide (ON) sequences were replaced with 5-benzofuran-modified 2′-deoxyuridine and uridine analogues. Depending on the position of modification the fluorescent nucleoside analogues distinguish antiparallel, mixed parallel-antiparallel and parallel stranded DNA and RNA GQ topologies from corresponding duplexes with significant enhancement in fluorescence intensity and quantum yield. Further, these GQ sensors enabled the development of a simple fluorescence binding assay to quantify topology- and nucleic acid-specific binding of small molecule ligands to GQ structures. Together, our results demonstrate that these nucleoside analogues are useful GQ probes, which are anticipated to provide new opportunities to study and discover efficient G-quadruplex binders of therapeutic potential. PMID:26202965

  18. A novel homogenous detection method based on the self-assembled DNAzyme labeled DNA probes with SWNT conjugates and its application in detecting pathogen.

    Science.gov (United States)

    Ding, Xinghua; Li, Hua; Deng, Le; Peng, Zhihui; Chen, Hui; Wang, Dan

    2011-07-15

    In this paper, a novel and cost-effective homogeneous detection method was constructed for the detection of genomic DNA and Staphylococcus aureus (S. aureus), based on the noncovalent assembly of DNAzyme-labeled detection probe and single-walled carbon nanotubes (SWNTs). When the target genomic DNA and hemin was existed in the detection solution, the detection probe wrapped on the SWNTs by π-stacking interactions would keep away from SWNTs and form a DNAzyme-self-assembly construction. This DNAzyme construction could catalyze 2,2'-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) (ABTS²⁻) and generate a colored product which could lead to the absorbance changes. Hence, according to its catalyzed capacity, the DNAzyme construction could amplify the detection signal. The concentration of target DNA could be quantified by exploiting their optical absorption changes at 414 nm and the concentration limit of detection of the method was 30 nM. And this detection method detected S. aureus quantitatively. In addition, this work proved that the method obtain higher detection sensitivity compared with the method without SWNTs because of the protection profile of SWNTs towards the detection probe.

  19. Comparison of the hydrophobic grid-membrane filter DNA probe method and the Health Protection Branch standard method for the detection of Listeria monocytogenes in foods.

    Science.gov (United States)

    Yan, W; Malik, M N; Peterkin, P I; Sharpe, A N

    1996-07-01

    The standard Health Protection Branch (HPB) method for the detection of L. monocytogenes in foods involves lengthy enrichment, selection and biochemical testing, requiring up to 8 days to complete. A hydrophobic grid-membrane filter (HGMF) method employing a digoxigenin-labelled listeriolysin O probe required 5 days to complete, and included an image-analysis system for electronic data acquisition. A total of 200 food samples encompassing 8 high-risk food groups (soft and semi-soft cheeses, packaged raw vegetables, frozen cooked shrimp, ground poultry, ground pork, ground beef, jellied meats, and pâté) were screened for the presence of L. monocytogenes by the two methods. Overall, 32 (16%) and 30 (15%) of the naturally-contaminated food samples tested positive for L. monocytogenes by the HPB and DNA methods, respectively. The DNA probe method was highly specific in discriminating L. monocytogenes from other Listeria spp. present in 50 of the samples tested. Results showed 94% sensitivity and 100% specificity between the two methods. The HGMF DNA probe method is an efficient and reliable alternative to the HPB standard method for detecting L. monocytogenes in foods.

  20. Solution structures of 2 : 1 and 1 : 1 DNA polymerase-DNA complexes probed by ultracentrifugation and small-angle X-ray scattering

    Energy Technology Data Exchange (ETDEWEB)

    Tang, Kuo-Hsiang; Niebuhr, Marc; Aulabaugh, Ann; Tsai, Ming-Daw [OSU; (Wyeth); (SSRL)

    2008-03-25

    We report small-angle X-ray scattering (SAXS) and sedimentation velocity (SV) studies on the enzyme-DNA complexes of rat DNA polymerase β (Pol β) and African swine fever virus DNA polymerase X (ASFV Pol X) with one-nucleotide gapped DNA. The results indicated formation of a 2 : 1 Pol β-DNA complex, whereas only 1 : 1 Pol X-DNA complex was observed. Three-dimensional structural models for the 2 : 1 Pol β-DNA and 1 : 1 Pol X-DNA complexes were generated from the SAXS experimental data to correlate with the functions of the DNA polymerases. The former indicates interactions of the 8 kDa 5'-dRP lyase domain of the second Pol β molecule with the active site of the 1 : 1 Pol β-DNA complex, while the latter demonstrates how ASFV Pol X binds DNA in the absence of DNA-binding motif(s). As ASFV Pol X has no 5'-dRP lyase domain, it is reasonable not to form a 2 : 1 complex. Based on the enhanced activities of the 2 : 1 complex and the observation that the 8 kDa domain is not in an optimal configuration for the 5'-dRP lyase reaction in the crystal structures of the closed ternary enzyme-DNA-dNTP complexes, we propose that the asymmetric 2 : 1 Pol β-DNA complex enhances the function of Pol β.

  1. Solution Structures of 2 : 1 And 1 : 1 DNA Polymerase - DNA Complexes Probed By Ultracentrifugation And Small-Angle X-Ray Scattering

    Energy Technology Data Exchange (ETDEWEB)

    Tang, K.H.; /Ohio State U.; Niebuhr, M.; /SLAC, SSRL; Aulabaugh, A.; /Wyeth Res. Biophys., Pearl River; Tsai, M.D.; /Ohio State U. /SLAC, SSRL

    2009-04-30

    We report small-angle X-ray scattering (SAXS) and sedimentation velocity (SV) studies on the enzyme-DNA complexes of rat DNA polymerase {beta} (Pol {beta}) and African swine fever virus DNA polymerase X (ASFV Pol X) with one-nucleotide gapped DNA. The results indicated formation of a 2 : 1 Pol {beta}-DNA complex, whereas only 1 : 1 Pol X-DNA complex was observed. Three-dimensional structural models for the 2 : 1 Pol {beta}-DNA and 1 : 1 Pol X-DNA complexes were generated from the SAXS experimental data to correlate with the functions of the DNA polymerases. The former indicates interactions of the 8 kDa 5{prime}-dRP lyase domain of the second Pol {beta} molecule with the active site of the 1 : 1 Pol {beta}-DNA complex, while the latter demonstrates how ASFV Pol X binds DNA in the absence of DNA-binding motif(s). As ASFV Pol X has no 5{prime}-dRP lyase domain, it is reasonable not to form a 2 : 1 complex. Based on the enhanced activities of the 2 : 1 complex and the observation that the 8 kDa domain is not in an optimal configuration for the 5{prime}-dRP lyase reaction in the crystal structures of the closed ternary enzyme-DNA-dNTP complexes, we propose that the asymmetric 2 : 1 Pol {beta}-DNA complex enhances the function of Pol {beta}.

  2. Fiberoptic metal detector capable of profile detection

    Science.gov (United States)

    Hua, Wei-Shu; Hooks, Joshua R.; Erwin, Nicholas A.; Wu, Wen-Jong; Wang, Wei-Chih

    2011-04-01

    The purpose of this paper is to develop a novel ferromagnetic polymeric metal detector system by using a fiber-optic Mach-Zehnder interferometer with a newly developed ferromagnetic polymer as the magnetostrictive sensing device. This ferromagnetic polymeric metal detector system is simple to fabricate, small in size, and resistant to RF interference (which is common in typical electromagnetic type metal detectors). Metal detection is made possible by disrupting the magnetic flux density present on the magnetostrictive sensor. This paper discusses the magnetic properties of the ferromagnetic polymers. In addition, the preliminary results of successful sensing of different geometrical metal shapes will be discussed.

  3. Fiberoptic metal detector capable of profile detection.

    Science.gov (United States)

    Hua, Wei-Shu; Hooks, Joshua R; Erwin, Nicholas A; Wu, Wen-Jong; Wang, Wei-Chih

    2011-03-31

    The purpose of this paper is to develop a novel ferromagnetic polymeric metal detector system by using a fiber-optic Mach-Zehnder interferometer with a newly developed ferromagnetic polymer as the magnetostrictive sensing device. This ferromagnetic polymeric metal detector system is simple to fabricate, small in size, and resistant to RF interference (which is common in typical electromagnetic type metal detectors). Metal detection is made possible by disrupting the magnetic flux density present on the magnetostrictive sensor. This paper discusses the magnetic properties of the ferromagnetic polymers. In addition, the preliminary results of successful sensing of different geometrical metal shapes will be discussed.

  4. Adaptive Holographic Fiber-Optic Interferometer

    Science.gov (United States)

    Kozhevnikov, Nikolai M.; Lipovskaya, Margarita J.

    1990-04-01

    Interaction of phase-modulated light beams in photorefractive local inertial responce media was analysed. Interaction of this type allows to registrate phase-modulated signals adaptively under low frequency phase disturbtion. The experiments on multimode fiber-optic interferometer with demodulation element based on photorefractive bacteriorhodopsin-doped polimer film are described. As the writing of dynamic phase hologram is an inertial process the signal fluctuations with the frequencies up to 100 Hz can be canceled. The hologram efficiencies are enough to registrate high frequency phase shifts ~10-4 radn.

  5. Next-generation repeat-free FISH probes for DNA amplification in glioblastoma in vivo: Improving patient selection to MDM2-targeted inhibitors.

    Science.gov (United States)

    Brunelli, Matteo; Eccher, Albino; Cima, Luca; Trippini, Tobia; Pedron, Serena; Chilosi, Marco; Barbareschi, Mattia; Scarpa, Aldo; Pinna, Giampietro; Cabrini, Giulio; Pilotto, Sara; Carbognin, Luisa; Bria, Emilio; Tortora, Giampaolo; Fioravanzo, Adele; Schiavo, Nicola; Meglio, Mario; Sava, Teodoro; Belli, Laura; Martignoni, Guido; Ghimenton, Claudio

    2017-01-01

    A next-generation FISH probe mapping to the MDM2 locus-specific region has recently been designed. The level of MDM2 gene amplification (high versus low) may allow selection of patients for cancer treatment with MDM2 inhibitors and may predict their responsiveness. We investigated the spectrum of MDM2 gene alterations using the new probes in vivo after visualizing single neoplastic cells in situ from a series of glioblastomas. Signals from next-generation repeat-free FISH interphase probes were identified in tissue microarrays that included 3 spots for each of the 48 cases. The murine double minutes (MDM2)-specific DNA probe and the satellite enumeration probe for chromosome 12 were used. Three cases (6%) showed more than 25 signals (high gene amplification), and 7 (15%) showed 3-10 signals (gains); among these, 4 cases (8%) had an equal number of MDM2 and centromeric signals on chromosome 12 (polyploidy). Genomic heterogeneity was observed only in 3 cases with low gene amplification. In our series, 6% of glioblastomas exhibited high MDM2 amplification (in vivo) with a pattern related to the known double minutes/chromothripsis phenomenon (in situ), and only cases with low amplification showed genomic heterogeneity. We concluded that the rate of MDM2 gene amplification can be a useful predictive biomarker to improve patient selection.

  6. Multivariate reference technique for quantitative analysis of fiber-optic tissue Raman spectroscopy.

    Science.gov (United States)

    Bergholt, Mads Sylvest; Duraipandian, Shiyamala; Zheng, Wei; Huang, Zhiwei

    2013-12-03

    We report a novel method making use of multivariate reference signals of fused silica and sapphire Raman signals generated from a ball-lens fiber-optic Raman probe for quantitative analysis of in vivo tissue Raman measurements in real time. Partial least-squares (PLS) regression modeling is applied to extract the characteristic internal reference Raman signals (e.g., shoulder of the prominent fused silica boson peak (~130 cm(-1)); distinct sapphire ball-lens peaks (380, 417, 646, and 751 cm(-1))) from the ball-lens fiber-optic Raman probe for quantitative analysis of fiber-optic Raman spectroscopy. To evaluate the analytical value of this novel multivariate reference technique, a rapid Raman spectroscopy system coupled with a ball-lens fiber-optic Raman probe is used for in vivo oral tissue Raman measurements (n = 25 subjects) under 785 nm laser excitation powers ranging from 5 to 65 mW. An accurate linear relationship (R(2) = 0.981) with a root-mean-square error of cross validation (RMSECV) of 2.5 mW can be obtained for predicting the laser excitation power changes based on a leave-one-subject-out cross-validation, which is superior to the normal univariate reference method (RMSE = 6.2 mW). A root-mean-square error of prediction (RMSEP) of 2.4 mW (R(2) = 0.985) can also be achieved for laser power prediction in real time when we applied the multivariate method independently on the five new subjects (n = 166 spectra). We further apply the multivariate reference technique for quantitative analysis of gelatin tissue phantoms that gives rise to an RMSEP of ~2.0% (R(2) = 0.998) independent of laser excitation power variations. This work demonstrates that multivariate reference technique can be advantageously used to monitor and correct the variations of laser excitation power and fiber coupling efficiency in situ for standardizing the tissue Raman intensity to realize quantitative analysis of tissue Raman measurements in vivo, which is particularly appealing in

  7. Technical aspects of typing for HLA-DP alleles using allele-specific DNA in vitro amplification and sequence-specific oligonucleotide probes. Detection of single base mismatches

    DEFF Research Database (Denmark)

    Fugger, L; Morling, N; Ryder, L P

    1990-01-01

    The polymerase chain reaction (PCR) is an effective method for in vitro DNA amplification which combined with probing with synthetic oligonucleotides can be used for, e.g., HLA-typing. We have studied the technical aspects of HLA-DP typing with the technique. DNA from mononuclear nucleated cells ...... mismatches may be detected in the PCR and typing for HLA-DP gene variants, which differ for only one base, may be performed.......The polymerase chain reaction (PCR) is an effective method for in vitro DNA amplification which combined with probing with synthetic oligonucleotides can be used for, e.g., HLA-typing. We have studied the technical aspects of HLA-DP typing with the technique. DNA from mononuclear nucleated cells...... was extracted with either a simple salting out method or phenol/chloroform. Both DNAs could be readily used for PCR. The MgC2 concentration of the PCR buffer and the annealing temperature of the thermal cycle of the PCR were the two most important variables. The MgCl2 concentration and the temperature must...

  8. Detection of virulence factors in culturable Escherichia coli isolates from water samples by DNA probes and recovery of toxin-bearing strains in minimal o-nitrophenol-beta-D-galactopyranoside-4-methylumbelliferyl-beta-D-g luc uronide media.

    OpenAIRE

    Martins, M T; Rivera, I G; Clark, D. L.; Olson, B H

    1992-01-01

    A total of 449 Escherichia coli isolates in treated and raw water sources were submitted to DNA-DNA hybridization using seven different DNA probes to detect homology to sequences that code for Shiga-like toxins I and II; heat-stabile and heat-labile toxins, adherence factors EAF and eae, and the fimbrial antigen of entero-hemorrhagic E. coli. Fifty-nine (13%) of the isolates demonstrated homology with one or more specific DNA probes. More than 50% of the isolates in treated water were not rec...

  9. Non-viral gene delivery carrier of probe type host molecule --Interactions between DNA and β-cyclodextrin derivative complexes (Ⅰ)

    Institute of Scientific and Technical Information of China (English)

    2006-01-01

    A host type non-virus gene delivery carrier, phenanthroline-β-cyclodextrin derivative host molecule, was produced which can be used as molecular probe. Interactions between DZY-1 and DNA were investigated by electrophoresis assay. Hind III enzyme inhibition assay was carried out using DNA condensates induced by host molecules or host- guest molecule complexes to explore their ability to inhibit enzyme digestion. Micro-structure of DNA condensates induced by host molecules and host-guest molecule complexes was observed by scanning electron microscope (SEM). Our work indicates the delivery mechanism of DZY-1 used as a gene delivery carrier and also provides a method to design and produce non-virus gene delivery carriers.

  10. Preparing to perform an awake fiberoptic intubation.

    LENUS (Irish Health Repository)

    Walsh, M E

    2012-02-03

    Fiberoptically guided tracheal intubation represents one of the most important advances in airway management to occur in the past thirty years. Perhaps its most important role is in management of the anticipated difficult airway. This is a situation in which the dangers of encountering the life-threatening "can\\'t intubate, can\\'t ventilate" situation can be avoided by placement of an endotracheal tube while the patient is awake. Although skill at the procedure of endoscopy is obviously necessary in this setting, these authors hold that success or failure of the technique frequently depends on the adequacy of preparation. These measures include 1) pre-operative assessment of the patient; 2) careful explanation of what lies in store; 3) "setting the stage"; 4) preparing the equipment to be used; and 5) preparing the patient (antisialogue, sedation, application of topical anesthesia to the upper airway). If these preparatory measures are carried out meticulously, the likelihood of performing a successful and comfortable awake fiberoptic tracheal intubation is greatly increased.

  11. Identification of a Toluene-Degrading Bacterium from a Soil Sample through H218O DNA Stable Isotope Probing ▿†

    Science.gov (United States)

    Woods, Angela; Watwood, Maribeth; Schwartz, Egbert

    2011-01-01

    DNA stable isotope probing (DNA-SIP) with H218O was used to identify a toluene-degrading bacterium in soil amended with 48 ppm toluene. After quantification of toluene degradation rates in soil, DNA was extracted from soil incubated with H218O, H216O, H216O and 48 ppm toluene, or H218O and 48 ppm toluene. A single DNA band formed along a cesium chloride gradient after isopycnic centrifugation of extracts from soils incubated with H216O. With extracts from soils to which only H218O was added, two distinct DNA bands formed, while three bands formed when DNA extracted from soil incubated with both H218O and toluene was analyzed. We suggest that this third band formed because toluene does not contain any oxygen atoms and toluene-degrading organisms had to transfer oxygen atoms from H218O into metabolic intermediates to form nucleic acids de novo. We extracted the third DNA band and amplified a large fraction of the bacterial 16S rRNA gene. Direct sequencing of the PCR product obtained from the labeled DNA, as well as cloned 16S rRNA amplicons, identified a known toluene degrader, Rhodococcus jostii RHA1. A toluene-degrading bacterial strain was subsequently isolated from soil and shown to be Rhodococcus jostii RHA1. Finally, quantitative real-time PCR analysis showed that the abundance of the 16S rRNA gene of Rhodococcus jostii RHA1 increased in soil after toluene exposure but not in soils from which toluene was withheld. This study indicates that H218O DNA-SIP can be a useful method for identifying pollutant-degrading bacteria in soil. PMID:21742928

  12. Molecular cytogenetic analysis of the Appenine endemic cyprinid fish Squalius lucumonis and three other Italian leuciscines using chromosome banding and FISH with rDNA probes.

    Science.gov (United States)

    Rossi, Anna Rita; Milana, Valentina; Hett, Anne Kathrin; Tancioni, Lorenzo

    2012-12-01

    Karyotype and other chromosomal characteristics of the Appenine endemic cyprinid fish, Toscana stream chub Squalius lucumonis, were analysed using conventional banding and FISH with 45S and 5S rDNA probes. The diploid chromosome number (2n = 50) and karyotype characteristics including pericentromeric heterochromatic blocks and GC-rich CMA(3)-positive sites corresponding to both positive Ag-NORs and 45S rDNA loci on the short arms of a single medium-sized submetacentric chromosome pair were consistent with those found in most European leuciscine cyprinids. On other hand, 5S rDNA FISH in the Toscana stream chub and three other Italian leuciscines, S. squalus, Rutilus rubilio and Telestes muticellus, revealed a species-specific hybridization pattern, i.e. signals on four (S. lucumonis), three (S. squalus and R. rubilio) and two (T. muticellus) chromosome pairs. Whereas all the species shared the 5S rDNA loci on the largest subtelocentric chromosome pair, a "leuciscine" cytotaxonomic marker, S. lucumonis showed both classes of rDNA loci tandem aligned on the short arms of chromosome pair No. 12. The present findings suggest that the observed high variability of 5S rDNA loci provides a powerful tool for investigation of karyotype differentiation in karyologically conservative leuciscine fishes.

  13. Chemical shift as a probe of molecular interfaces: NMR studies of DNA binding by the three amino-terminal zinc finger domains from transcription factor IIIA

    Energy Technology Data Exchange (ETDEWEB)

    Foster, Mark P.; Wuttke, Deborah S.; Clemens, Karen R.; Jahnke, Wolfgang; Radhakrishnan, Ishwar; Tennant, Linda; Reymond, Martine; Chung, John; Wright, Peter E. [Scripps Research Institute, Department of Molecular Biology and Skaggs Institute for Chemical Biology (United States)

    1998-07-15

    We report the NMR resonance assignments for a macromolecular protein/DNA complex containing the three amino-terminal zinc fingers (92 amino acid residues) of Xenopus laevis TFIIIA (termed zf1-3) bound to the physiological DNA target (15 base pairs), and for the free DNA. Comparisons are made of the chemical shifts of protein backbone{sup 1} H{sup N}, {sup 15}N,{sup 13} C{sup {alpha}} and{sup 13} C{sup {beta}} and DNA base and sugar protons of the free and bound species. Chemical shift changes are analyzed in the context of the structures of the zf1-3/DNA complex to assess the utility of chemical shift change as a probe of molecular interfaces. Chemical shift perturbations that occur upon binding in the zf1-3/DNA complex do not correspond directly to the structural interface, but rather arise from a number of direct and indirect structural and dynamic effects.

  14. MGB probe assay for rapid detection of mtDNA11778 mutation in the Chinese LHON patients by real-time PCR

    Institute of Scientific and Technical Information of China (English)

    Jian-yong WANG; Yang-shun GU; Jing WANG; Yi TONG; Ying WANG; Jun-bing SHAO; Ming QI

    2008-01-01

    Objective:Leber's hereditary optic neuropathY (LHON)is a maternally inherited degeneration of the optic nerve caused by point mutations of mitochondrial DNA(mtDNA).Many unsolved questions regarding the penetrance and pathophysiological mechanism of LHON demand efficient and reliable mutation testing.This study aims to develop a minor groove binder(MGB) probe assay for rapid detection of mtDNA11778 mutation and heteroplasmy in Chinese LHON patients by real-time polymerase chain reaction(PCR).Methods:Forty-eight patients suspected of having LHON and their maternal relatives underwent a molecular genetic evaluation,with 20 normal individuals as a control group at the same time.A real-time PCR involving two MGB probes was used to detect the mtDNA 11778 mutation and heteroplasmy.A linear standard curve was obtained by pUCmLHONG and pUCmLHONA clones.Results:All 48 LHON patients and their matemal relatives were positive for mtDNA 11778 mutation in our assay,27 heteroplasmic and 21 homoplasmic.Eighteen cases did not show an occurrence of the disease,while 9 developed the disease among the 27 heteroplasmic mutation cases.Eleven did not show an occurrence of the disease,while 10 cases developed the disease among 21 homoplasmic mutation cases.There was a significant difierence in the incidence between the heteroplasmic and the homoplasmic mutation types.The time needed for running a real-time PCR assay was only 80 min.Conclusion:This real-time PCR assay is a rapid,reliable method for mtDNA mutation detection as well as heteroplasmy quantification.Detecting this ratio is very important for predicting phenotypic expression of unaffected carriers.

  15. Immunogold-silver staining (IGSS) based U-bent fiberoptic sandwich biosensor

    Science.gov (United States)

    Ramakrishna, B.; Sai, V. V. R.

    2016-11-01

    An evanescent wave absorbance (EWA) based U-bent fiberoptic sandwich immunobiosensor with human IgG detection limits of 6.67 fM (1 pg/ml of human IgG) and 66.7 aM (10 fg/ml of HIgG) is demonstrated by exploiting immunogold labels and subsequently silver enhancement respectively. Such very low detection limits were achieved with the help of enhanced evanescent filed at the bend region of U-bent optical fiber probe that allows efficient interaction of light with 40 nm immunogold labels on the probe surface resulting in measurable optical absorbance changes. The other significant advantages of the demonstrated sensing scheme are low cost optoelectronic instrumentation consisting of an commercial green LED and a photodetector (S150C, Thorlabs Inc.), small volumes of sample and immunogold reagent each of 25 μl and rapid detection in 20 min. These results from the plasmonic fiberoptic biosensor demonstrate its huge potential for development of point-of-care diagnostic devices for sensitive and rapid detection of analytes.

  16. Development of a polymer based fiberoptic magnetostrictive metal detector system.

    Science.gov (United States)

    Hua, Wei Shu; Hooks, Joshua Rosenberg; Wu, Wen Jong; Wang, Wei Chih

    2010-10-01

    This paper presents a new metal detector using a fiberoptic magnetostriction sensor. The metal sensor uses a fiber-optic Mach-Zehnder interferometer with a newly developed ferromagnetic polymer as the magnetostrictive sensing material. This polymeric magnetostrictive fiberoptic metal sensor is simple to fabricate, small in size, and resistant to RF interference (which is common in typical electromagnetic type metal detectors). Metal detection is based on disruption of the magnetic flux density across the magnetostriction sensor. In this paper, characteristics of the material being sensed and magnetic properties of the ferromagnetic polymers will be discussed.

  17. Fiber-Optic Chemical Sensors and Fiber-Optic Bio-Sensors

    Directory of Open Access Journals (Sweden)

    Marie Pospíšilová

    2015-09-01

    Full Text Available This review summarizes principles and current stage of development of fiber-optic chemical sensors (FOCS and biosensors (FOBS. Fiber optic sensor (FOS systems use the ability of optical fibers (OF to guide the light in the spectral range from ultraviolet (UV (180 nm up to middle infrared (IR (10 μm and modulation of guided light by the parameters of the surrounding environment of the OF core. The introduction of OF in the sensor systems has brought advantages such as measurement in flammable and explosive environments, immunity to electrical noises, miniaturization, geometrical flexibility, measurement of small sample volumes, remote sensing in inaccessible sites or harsh environments and multi-sensing. The review comprises briefly the theory of OF elaborated for sensors, techniques of fabrications and analytical results reached with fiber-optic chemical and biological sensors.

  18. Comparison of Xpert MTB/RIF Assay and GenoType MTBDRplus DNA Probes for Detection of Mutations Associated with Rifampicin Resistance in Mycobacterium tuberculosis.

    Directory of Open Access Journals (Sweden)

    Arfatur Rahman

    Full Text Available GeneXpert MTB/RIF (Xpert and Genotype MTBDRplus (DRplus are two World Health Organization (WHO endorsed probe based molecular drug susceptibility testing (DST methods for rapid diagnosis of drug resistant tuberculosis. Both methods target the same 81 bp Rifampicin Resistance Determining Region (RRDR of bacterial RNA polymerase β subunit (rpoB for detection of Rifampicin (RIF resistance associated mutations using DNA probes. So there is a correspondence of the probes of each other and expected similarity of probe binding.We analyzed 92 sputum specimens by Xpert, DRplus and LJ proportion method (LJ-DST. We compared molecular DSTs with gold standard LJ-DST. We wanted to see the agreement level of two molecular methods for detection of RIF resistance associated mutations. The 81bp RRDR region of rpoB gene of discrepant cases between the two molecular methods was sequenced by Sanger sequencing.The agreement of Xpert and DRplus with LJ-DST for detection of RIF susceptibility was found to be 93.5% and 92.4%, respectively. We also found 92.4% overall agreement of two molecular methods for the detection of RIF susceptibility. A total of 84 out of 92 samples (91.3% had agreement on the molecular locus of RRDR mutation by DRplus and Xpert. Sanger sequencing of 81bp RRDR revealed that Xpert probes detected seven of eight discrepant cases correctly and DRplus was erroneous in all the eight cases.Although the overall concordance with LJ-DST was similar for both Xpert and DRplus assay, Xpert demonstrated more accuracy in the detection of RIF susceptibility for discrepant isolates compared with DRplus. This observation would be helpful for the improvement of probe based detection of drug resistance associated mutations especially rpoB mutation in M. tuberculosis.

  19. Synthesis of circular double-stranded DNA having single-stranded recognition sequence as molecular-physical probe for nucleic acid hybridization detection based on atomic force microscopy imaging.

    Science.gov (United States)

    Nakano, Koji; Matsunaga, Hideshi; Murata, Masaharu; Soh, Nobuaki; Imato, Toshihiko

    2009-08-01

    A new class of DNA probes having a mechanically detectable tag is reported. The DNA probe, which consists of a single-stranded recognition sequence and a double-stranded circular DNA entity, was prepared by polymerase reaction. M13mp18 single strand and a 32mer oligodeoxynucleotide whose 5'-end is decorated with the recognition sequence were used in combination as template and primer, respectively. We have successfully demonstrated that the DNA probe is useful for bioanalytical purposes: by deliberately attaching target DNA molecules onto Au(111) substrates and by mechanically reading out the tag-entity using a high-resolution microscopy including atomic force microscopy, visualization/detection of the individual target/probe DNA conjugate was possible simply yet straightforwardly. The present DNA probe can be characterized as a 100%-nucleic acid product material. It is simply available by one-pod synthesis. A surface topology parameter, image roughness, has witnessed its importance as a quantitative analysis index with particular usability in the present visualization/detection method.

  20. The commonly-used DNA probe for diffusely-adherent Escherichia coli cross-reacts with a subset of enteroaggregative E. coli

    Directory of Open Access Journals (Sweden)

    Fletcher Jonathan N

    2009-12-01

    Full Text Available Abstract Background The roles of diffusely-adherent Escherichia coli (DAEC and enteroaggregative E. coli (EAEC in disease are not well understood, in part because of the limitations of diagnostic tests for each of these categories of diarrhoea-causing E. coli. A HEp-2 adherence assay is the Gold Standard for detecting both EAEC and DAEC but DNA probes with limited sensitivity are also employed. Results We demonstrate that the daaC probe, conventionally used to detect DAEC, cross-reacts with a subset of strains belonging to the EAEC category. The cross hybridization is due to 84% identity, at the nucleotide level, between the daaC locus and the aggregative adherence fimbriae II cluster gene, aafC, present in some EAEC strains. Because aaf-positive EAEC show a better association with diarrhoea than other EAEC, this specific cross-hybridization may have contributed to an over-estimation of the association of daaC with disease in some studies. We have developed a discriminatory PCR-RFLP protocol to delineate EAEC strains detected by the daaC probe in molecular epidemiological studies. Conclusions A PCR-RFLP protocol described herein can be used to identify aaf-positive EAEC and daaC-positive DAEC and to delineate these two types of diarrhoeagenic E. coli, which both react with the daaC probe. This should help to improve current understanding and future investigations of DAEC and EAEC epidemiology.

  1. Detection of Human Papillomavirus DNA by AffiProbe HPV-DNA Test Kit in Cervical Scrapes or Biopsies-Histopathologic Correlates

    OpenAIRE

    Pekka Nieminen; Tarja Jalava; Arja Kallio; Marjut Ranki; Jorma Paavonen

    1994-01-01

    Objective: The aim of this study was to evaluate and compare the efficacy of punch biopsies and cervical scrapes in the detection of human papillomavirus (HPV) DNA from the cervix and compare the results with the histopathologic diagnosis. Methods: The specimens were collected simultaneously, and HPV DNA was detected using a liquid hybridization test. Results: Biopsies and scrapes were equally efficient, but each detected only two-thirds of all HPV-DNA-positive patients. Thus, the positivity ...

  2. 两种念珠菌DNA探针的研制及应用%Preparation and using of two kinds of Candida DNA probe

    Institute of Scientific and Technical Information of China (English)

    佘菲菲; 朱苹; 陈月秀; 沈建箴; 吕联煌

    2001-01-01

    目的 制备两种念珠菌DNA探针并应用于临床诊断。方法 对聚合酶链反应扩增的白色念珠菌标准株的E03基因的125bp片段,用半抗原地高辛标记;合成仪合成的E03基因中25bp特异寡核苷酸,用生物素标记。检测两种探针显色敏感度,并与23株临床分离的白色念珠菌、热带念珠菌,近平滑念珠菌、克柔念珠菌、星形念珠菌及大肠埃希菌等细菌进行斑点杂交试验。结果 Dig-125bpDNA探针显色敏感度为0.01pg,仅与白色念珠菌杂交,且对临床分离株检测的结果与传统厚膜孢子鉴定法相符。Bio-25bpDNA探针显色敏感度为20pg,与白色念珠菌、热带念珠菌、近平滑念珠菌及星形念珠菌杂交。结论 Bio-25bpDNA探针可用于念珠菌初步鉴定,而Dig-125bpDNA探针可用于白色念珠菌的鉴定。%Objective Preparation and using of Candida DNA probe.Methods The125 bp DNA from E03 gene of C.albicans was amplified by PCR,then labeled by hepen digoxigenin,and the 25 bp specific sequence from E03 gene was synthesized and labeled with biotin.The sensitivity of these probes was detected,and the specificity was detected by dot-blotting hybridization assay with 23 strains C.albicans isolated from patients,C.paropsilosis,C.tropicalis,C.krusei,C.stellatoidea and E.coli and so on.Results The Dig-125 bp DNA probe,only hybridized with C.albicans, whose sensitivity of color development was 0.01 pg,and the hybridization result of the isolated strains was coincidental with the result of traditional identification for C.albicans.The Bio-25 bp DNA probe,hybridized with C.albicans,C.paropsilosis,C.tropicalis and C.stellatoidea,whose sensitivity of color development was 20 pg.Conclusion The Dig-125 bp DNA probe is specific and sensitive to C.albicans and suitable for the indentification of clinical C.albicans infection,while the Bio-25 bp DNA probe is suitable for the indentification of clinical candida infection.

  3. PCR associated with hybridization with DNA radioactive probes for diagnosis of asymptomatic infection caused by Leishmania Chagasi; PCR associado a hibridizacao com sondas radioativas de DNA para a identificacao de infeccao subclinica causada por Leishmania Chagasi

    Energy Technology Data Exchange (ETDEWEB)

    Andrade, Antero Silva Ribeiro de [Centro de Desenvolvimento da Tecnologia Nuclear (CDTN), Belo Horizonte, MG (Brazil); Moreno, Elizabeth Castro [Fundacao Nacional de Saude, Belo Horizonte, MG (Brazil). Coordenacao Regional de Minas Gerais; Gomes, Rosangela Fatima; Melo, Maria Norma de; Carneiro, Mariangela [Minas Gerais Univ., Belo Horizonte, MG (Brazil). Dept. de Parasitologia; Fernandes, Octavio [Fundacao Inst. Oswaldo Cruz (FIOCRUZ), Rio de Janeiro, RJ (Brazil). Dept. de Medicina Tropical

    2002-07-01

    Detection systems for diagnosis of leishmaniasis based on PCR are very promising due to their sensitivity and specificity. Secondary detection by specific radioactive DNA probes, able to type the PCR amplified products, increase the specificity and raise about tem-fold the sensitivity of the assay. The aim of this work was evaluate PCR and hybridization as a tool to identify Leishmania (Leishmania) chagasi (the specie that cause the visceral leishmaniasis in Brazil) infection in asymptomatic persons living in a endemic area. Material and Methods: A group of 226 asymptomatic individuals, living in General Carneiro (MG), was selected. Blood samples were harvested and the DNA extracted from the mononucleate cells. PCR was performed using primers addressed to the kinetoplast DNA minicircles. This protocol gives a positive reaction for all Leishmania species. The amplified products were further hybridized with cloned L.chagasi minicircles labeled with {sup 32} P. Results: were identified 111 samples PCR positive, 2 of them hybridization negative and 133 samples hybridization positive, 24 of them PCR negative. The occurrence of samples with hybridization positive and PCR negative was expected since hybridization, with DNA probes labeled with {sup 32} P, increase the sensitivity of the assay. The samples that presented positive PCR and negative hybridization were probably due the presence of other Leishmania species, likely L. (V.) braziliensis (that produce tegumentary leishmaniasis in the region), since L. (L.) chagasi cloned minicircles were used as hybridization probe. We conclude that this procedure is a valuable tool to access subclinical L. (L.) chagasi infections in epidemiological studies. (author)

  4. Methylation at the PW71 locus on chromosome 15 in DNA derived from CVS and from amniocytes; implications for the use of the PW71 probe in prenatal diagnosis of the Prader-Willi and Angleman syndromes

    Energy Technology Data Exchange (ETDEWEB)

    Telleria, P.; Yu, C.C.; Brown, S. [Columbia Univ., New York, NY (United States)

    1994-09-01

    The probe PW71 spans a HpaII site in the Prader-Willi/Angleman Syndrome critical region on chromosome 15. A single Southern blot with this probe can be used to detect deletion and uniparental disomy. We attempted to determine the methylation state of the PW71 locus in DNA derived from prenatal sources. Southern blots of HindIII and HindIII/HpaII double digests of DNA from cultured amniocytes and CVS specimens were prepared and probed with the PW71 probe. The results from 6 cultured CVS specimens indicate that several HPAII sites recognized by the PW71 probe are not methylated in trophoblast. Four amniotic fluid cultures gave results which were not different from lymphocyte-derived DNA; however, in several cases, amniotic fluid cultures resulted in Southern blots identical to those from CVS. Since we did not have verified prenatal cases of chromosome 15 uniparental disomy, we were unable to determine whether the parent-of-origin specific methylation present in lymphocyte DNA is also present in amniocyte DNA. We conclude that prenatal determination of chromosome 15 uniparental disomy with this probe will be unreliable.

  5. Diagnosis of canine visceral leishmaniasis with radiolabelled probes: comparison of the kDNA PCR-hybridization with three molecular methods in different clinical samples

    Energy Technology Data Exchange (ETDEWEB)

    Ferreira, Aline Leandra C.; Ferreira, Sidney A.; Carregal, Virginia M.; Andrade, Antero Silva R., E-mail: antero@cdtn.br [Centro de Desenvolvimento da Tecnologia Nuclear (CDTN/CNEN-MG), Belo Horizonte, MG (Brazil). Lab. de Radiobiologia; Melo, Maria N., E-mail: melo@icb.ufmg.br [Departamento de Parasitologia. Instituto de Ciencias Biologicas. Universidade Federal de Minas Gerais, Belo Horizonte, MG (Brazil)

    2011-07-01

    Leishmania (Leishmania) chagasi is responsible for visceral leishmaniasis (VL) in Brazil and the dog is the main domestic reservoir. Disease control is based on the elimination of infected animals and the use of a sensitive and specific diagnostic test is necessary. The Brazilian VL control program emphasizes serologic surveys, mainly using the enzyme-linked immunosorbent assay (ELISA) and the immunofluorescence antibody test (IFAT), followed by the elimination of the seropositive dogs. However, these techniques present limitations in terms of sensitivity and specificity. The Polymerase Chain Reaction (PCR) associated to hybridization with DNA probes labeled with {sup 32}P has been recognized as a valuable tool for Leishmania identification. In this study, the sensitivity of kDNA PCR hybridization method was compared with three other molecular methods: Internal Transcribed Spacer 1 Nested PCR (ITS-1nPCR), Leishmania nested PCR (LnPCR) and Seminested kDNA PCR (kDNA snPCR). The comparison was performed in different clinical specimens: conjunctival swab, skin, blood and bone marrow. A group of thirty symptomatic dogs, positive in the parasitological and serological tests, was used. When. The techniques targeting kDNA mini-circles (kDNA snPCR and KDNA PCR-hybridization) showed the worst result for blood samples. The KDNA-PCR hybridization showed the best sensitivity for conjunctival swab. By comparing the samples on the basis of positivity obtained by the sum of all methods, the blood showed the worst outcome (71/120).The bone marrow showed the highest positivity (106/120), followed by conjunctival swab (100/120) and skin (89/120). Since the bone marrow samples are unsuitable for routine epidemiological surveys, the conjunctival swab was recommended because it allows high sensitivity, especially when associated with kDNA PCR hybridization method, and is a noninvasive sampling method. (author)

  6. Probing radiation damage by alternated current conductivity as a method to characterize electron hopping conduction in DNA molecules

    Energy Technology Data Exchange (ETDEWEB)

    Gomes, Paulo J.; Coelho, Margarida; Antonio Ribeiro, Paulo; Raposo, Maria [CEFITEC, Departamento de Fisica, Faculdade de Ciencias e Tecnologia, FCT, Universidade Nova de Lisboa, 2829-516 Caparica (Portugal); Dionisio, Madalena [REQUIMTE, Departamento de Quimica, Faculdade de Ciencias e Tecnologia, FCT, Universidade Nova de Lisboa, 2829-516 Caparica (Portugal)

    2012-09-17

    Analysis of AC electrical conductivity of deoxyribonucleic acid (DNA) thin films, irradiated with ultraviolet (UV) light, revealed that electrical conduction arises from DNA chain electron hopping between base-pairs and phosphate groups. The hopping distance calculated from correlated barrier hopping model equals the distance between DNA base-pairs, which is consistent with the loss of conductivity with irradiation time arising from a decrease in phosphates groups. In the high frequency regime, at a given frequency, real part of conductivity strongly depends on irradiation time particularly for low dose levels suggesting the use of DNA based films for UV radiation sensors.

  7. Detection of Listeria monocytogenes by direct colony hybridization on hydrophobic grid-membrane filters by using a chromogen-labeled DNA probe.

    Science.gov (United States)

    Peterkin, P I; Idziak, E S; Sharpe, A N

    1991-02-01

    A DNA probe specific for Listeria monocytogenes was isolated from a beta-hemolytic recombinant clone of an L. monocytogenes gene bank. It was labeled with horseradish peroxidase and used in a direct colony hybridization method on hydrophobic grid-membrane filters for the detection of the organism. Following color development of the chromogen, a commercial counter (HGMF Interpreter) was able to detect and count the organisms electronically. The method gave a positive reaction with 70 L. monocytogenes strains, while showing a negative reaction with 10 strains of other Listeria spp. and with 20 organisms of other genera.

  8. Probing the dynamic differential stiffness of dsDNA interacting with RecA in the enthalpic regime.

    Science.gov (United States)

    Lien, Chia-Hui; Wei, Ming-Tzo; Tseng, Te-Yu; Lee, Chien-Der; Wang, Chung; Wang, Ting-Fang; Ou-Yang, H Daniel; Chiou, Arthur

    2009-10-26

    RecA plays a central role in homologous recombination of DNA. When RecA combines with dsDNA to form RecA-dsDNA nucleofilament, it unwinds dsDNA and changes its structure. The unwinding length extension of a DNA segment interacting with RecA has been studied by various techniques, but the dynamic differential stiffness of dsDNA conjugating with RecA has not been well characterized. We applied oscillatory optical tweezers to measure the differential stiffness of dsDNA molecules, interacting with RecA, as a function of time at a constant stretching force of 33.6pN. The values of the differential stiffness of DNA (for stretching force in the range of 20.0pN to 33.6pN) measured by oscillatory optical tweezers, both before and after its interaction with RecA, are consistent with those measured by stationary optical tweezers. In the dynamic measurement, we have shown that the association (or binding) rate increases with higher concentration of RecA; besides, we have also monitored in real-time the dissociation of RecA from the stretched RecA-dsDNA filament as ATPgammaS was washed off from the sample chamber. Finally, we verified that RecA (I26C), a form of RecA mutant, does not affect the differential stiffness of the stretched DNA sample. It implies that mutant RecA (I26C) does not bind to the DNA, which is consistent with the result obtained by conventional biochemical approach.

  9. Comparison of the stem-loop and linear probe-based electrochemical DNA sensors by alternating current voltammetry and cyclic voltammetry.

    Science.gov (United States)

    Yang, Weiwei; Lai, Rebecca Y

    2011-12-06

    Here we systematically characterized the sensor performance of the stem-loop probe (SLP) and linear probe (LP) electrochemical DNA sensors using alternating current voltammetry (ACV) and cyclic voltammetry (CV), with the goal of generating the set of operational criteria that best suits each sensor architecture, in addition to elucidating the signaling mechanism behind these sensors. Although the LP sensor shows slightly better % signal suppression (SS) upon hybridization with the perfect match target at 10 Hz, our frequency-dependent study suggests that it shows optimal % SS only in a very limited AC frequency range. Similar results are observed in CV studies in which the LP sensor, when compared to the SLP sensor, displays a narrower range of voltammetric scan rates where the optimal % SS can be achieved. More importantly, the difference between the two sensors' performance is particularly pronounced if the change in integrated charge (Q) upon target hybridization, rather than the peak current (I), is measured in CV. The temperature-dependent study further highlights the differences between the two sensors, where the LP sensor, owing to the flexible linear probe architecture, is more readily perturbed by temperature changes. Both SLP and LP sensors, however, show a loss of % SS when operated at elevated temperatures, despite the significant improvement in the hybridization kinetics. In conjunction with the ACV, CV, and temperature-dependent studies, the electron-transfer kinetics study provides further evidence in support of the proposed signaling mechanism of these two sensors, in which the SLP sensor's signaling efficiency and sensor performance is directly linked to the hybridization-induced conformational change in the redox-labeled probe, whereas the performance of the LP sensor relies on the hybridization-induced change in probe dynamics. © 2011 American Chemical Society

  10. Genotyping of velvet antlers for identification of country of origin using mitochondrial DNA and fluorescence melting curve analysis with locked nucleic acid probes.

    Science.gov (United States)

    Ahn, Jeong Jin; Kim, Youngjoo; Hong, Ji Young; Kim, Gi Won; Hwang, Seung Yong

    2016-07-01

    Velvet antlers are used medicinally in Asia and possess various therapeutic effects. Prices are set according to the country of origin, which is unidentifiable to the naked eye, and therefore counterfeiting is prevalent. Additionally, antlers of the Canadian elk, which can generate chronic wasting disease, are prevalently smuggled and distributed in the market. Thus, a method for identifying the country of origin of velvet antlers was developed, using polymorphisms in mitochondrial DNA, fluorescence melting curve analysis and analysis of locked nucleic acids (LNA). This combined method is capable of identifying five genotypes of velvet antlers in a single experiment using two probes. It also has advantages in multiplexing, simplicity and efficiency in genotyping, when compared to real-time PCR or microarrays. The developed method can be used to improve identification rates in the velvet antler market and, by extension, research based on polymorphisms in DNA sequences.

  11. DNA Probe and Cleavage Studies Based on the Interaction between DNA and Yb, Er Complexes of 2-[(Trifluoroaceto)aceto]thinophene-piperidine Ligands

    Institute of Scientific and Technical Information of China (English)

    YIN Caixia; HUO Fangjun; WU Yanbo; LIU Yanlin; YANG Pin

    2009-01-01

    Two kinds of Ln complexes of [Ln(TTA)4]·Hp (Ln=Yb or Er, TrA=2-[(Trifluoroaceto)aceto]thinophene, HP =piperidine) have been synthesized and characterized, and their DNA-binding properties investigated using UV spectra, fluorescent spectra, viscometry and molecular modeling. The results show that they can intercalate into the double helices of DNA. More important thing is that their fluorescence intensity can be enhanced by DNA, thererfore, a sensitive fluorescence method for the determination of DNA may be developed. The cleavage reaction on plasmid DNA has been monitored by agarose gel electrophoresis. Interestingly, the complexes can cleave circular plasmid pBR322 DNA at pH=7.2 and 37 ℃. In addition, BDNPP [bis(2,4-dinitropheny1)-phosphate] was chosen as a model compound to further study their cleavage mechanism of pBR322 DNA. From the first-order kinetics equation, it was proved indirectly that the mechanism may be a hydrolytic cleavage.

  12. Tuning two-photon photoluminescence of gold nanoparticle aggregates with DNA and its application as turn-on photoluminescence probe for DNA sequence detection.

    Science.gov (United States)

    Yuan, Peiyan; Ma, Rizhao; Guan, Zhenping; Gao, Nengyue; Xu, Qing-Hua

    2014-08-13

    Plasmon coupling between noble metal nanoparticles has been known to dramatically enhance linear and nonlinear optical properties of nearby chromophores and metal nanoparticles themselves. The interparticle distance is expected to have significant influence on the coupling strength. Here we have prepared DNA tuned Au nanoparticle assemblies with well controlled separation distances from 2.0 to 12.2 nm to investigate plasmon coupling strength and particle size effects on two-photon photoluminescence (TPPL) enhancement. TPPL intensities of these DNA coupled nanoassemblies were found to increase rapidly as the separation distance decreases. The largest TPPL enhancement factors of 115 and 265 were achieved at the shortest available separation distance of 2.0 nm for 21 and 41 nm Au NPs-dsDNA assemblies, respectively. We have further utilized DNA induced coupling of Au NPs and TPPL enhancement to develop a two-photon sensing scheme for detection of DNA sequences. This TPPL based method displayed high sensitivity with a limit of detection of 2.9 pM and excellent selectivity against ssDNA with mismatched bases. A single mismatch can be easily differentiated at room temperature. Taking the unique advantages of two-photon excitation, this method could be potentially further extended to DNA detection inside cells or even in vivo. These findings can provide important insight for fundamental understanding of plasmon-coupling enhanced TPPL and development of various two-photon excitation based applications.

  13. Isolate-Specific Detection of Grapevine fanleaf virus from Xiphinema index Through DNA-Based Molecular Probes.

    Science.gov (United States)

    Finetti-Sialer, M M; Ciancio, A

    2005-03-01

    ABSTRACT Tests with a real-time reverse transcription-polymerase chain reaction (RT-PCR) were performed on specimens of Xiphinema index collected from the rhizosphere of Grapevine fanleaf virus (GFLV)-infected grapevines at Palagiano, Italy. A 1,157-bp fragment of the GFLV RNA-2 coat protein (CP) gene was amplified and sequenced. A fluorescent Scorpion probe was designed to detect a highly conserved CP region. A second region with isolate-specific multiple nucleotide polymorphisms was used to detect GFLV isolates using molecular beacons (MB). The Scorpion probe allowed quantitative estimation of GFLV RNA-2 in single nematodes, using a dilution series of a 692-nucleotide transcript of the CP gene. The assay allowed detection of GFLV RNA-2 in individual X. index, with a minimum template threshold of 800 fg or 2.8 x 10(6) RNA-2 molecules per nematode. The CP fragment used for GFLV detection with the Scorpion probe appeared highly conserved among isolates. The probes were tested against other GFLV isolates, which were recognized by the species-specific Scorpion probe and by the corresponding MB specific to the particular isolate. Both tests appeared useful as diagnostic tools or for studies on GFLV in acquisition, retention, and transmission experiments.

  14. Thermodynamic and structural study of pyrene-1-carboxaldehyde/DNA interactions by molecular spectroscopy: Probing intercalation and binding properties

    Energy Technology Data Exchange (ETDEWEB)

    Grueso, E. [Department of Physical Chemistry, University of Seville, C/Profesor Garcia Gonzalez s/n, 41012 Seville (Spain); Prado-Gotor, R., E-mail: pradogotor@us.es [Department of Physical Chemistry, University of Seville, C/Profesor Garcia Gonzalez s/n, 41012 Seville (Spain)

    2010-08-03

    Graphical abstract: The exocyclic carbonyl compound pyren-1-carboxyaldehyde, (1-PyCHO), binds to the ctDNA in an intercalative mode. Two possible angular orientations for intercalation into base-pairs are possible. Induced circular dichroism measurements indicate that the intercalation orientation of 1-PyCHO into DNA could be heterogeneous, that is, multiple binding orientations of the pyren-1-carboxyaldehyde must be involved. - Abstract: The binding of pyrene-1-carboxaldehyde (1-PyCHO) with ctDNA was investigated through absorption, intrinsic and induced circular dichroism, viscosity measurements and steady-state fluorescence. The binding and the number of monomer units of the polymer involved in the binding of one dye molecule (site size) have been quantified. The results indicated that the 1-PyCHO molecule binds to the ctDNA in an intercalative mode. The spectroscopic evidence of this intercalation process is also corroborated by the effect of urea, iodide-induced fluorescence quenching of pyrene-1-carboxaldehyde and competitive binding using a fluorescent intercalator, SYBR Green I (SG). The induced circular dichroism (ICD) spectra of pyrene-1-carboxaldehyde complexed with ctDNA show that pyrene-1-carboxaldehyde intercalates into ctDNA and that the intercalation orientation of pyrene to the DNA base-pairs long axis is heterogeneous. On the other hand, the intrinsic circular dichroism (CD) spectra show a stabilization of the right-handed B form of ctDNA, due to the intercalation process.

  15. Flow-Induced Dispersion Analysis for Probing Anti-dsDNA Antibody Binding Heterogeneity in Systemic Lupus Erythematosus Patients

    DEFF Research Database (Denmark)

    Poulsen, Nicklas N; Pedersen, Morten E; Østergaard, Jesper

    2016-01-01

    Detection of immune responses is important in the diagnosis of many diseases. For example, the detection of circulating autoantibodies against double-stranded DNA (dsDNA) is used in the diagnosis of Systemic Lupus Erythematosus (SLE). It is, however, difficult to reach satisfactory sensitivity...

  16. Probing the Relationship between Anti-Pneumocystis carinii Activity and DNA Binding of Bisamidines by Molecular Dynamics Simulations

    Directory of Open Access Journals (Sweden)

    Teresa Żołek

    2015-04-01

    Full Text Available The anti-Pneumocystis carinii activity of 13 synthetic pentamidine analogs was analyzed. The experimental differences in melting points of DNA dodecamer 5'-(CGCGAATTCGCG2-3' complexes (ΔTm, and in the biological activity measured using ATP bioluminescence assay (IC50 together with the theoretical free energy of DNA-ligand binding estimated by the proposed computational protocol, showed that the experimental activity of the tested pentamidines appeared to be due to the binding to the DNA minor groove with extended AT sequences. The effect of heteroatoms in the aliphatic linker, and the sulfonamide or methoxy substituents on the compound inducing changes in the interactions with the DNA minor groove was examined and was correlated with biological activity. In computational analysis, the explicit solvent approximation with the discrete water molecules was taken into account, and the role of water molecules in the DNA-ligand complexes was defined.

  17. Fluorescent probes based on side-chain chlorinated benzo[a]phenoxazinium chlorides: Studies of interaction with DNA

    Science.gov (United States)

    Raju, B. Rama; Gonçalves, M. Sameiro T.; Coutinho, Paulo J. G.

    2017-01-01

    The interaction of DNA with six water soluble benzo[a]phenoxazinium chlorides mono- or di-substituted with 3-chloropropyl groups at the O and N of 2- and 9-positions, along with methyl, hydroxyl and amine terminal groups at 5-positions, was investigated by photophysical techniques. The results indicated that almost all compounds intercalated in DNA base pairs at phosphate to dye ratio higher than 5. At lower values of this ratio, electrostatic binding mode with DNA was observed. Groove binding was detected mainly for the benzo[a]phenoxazinium dye with NH2·HBr terminal. The set of six benzo[a]phenoxazinium chlorides proved successful to label the migrating DNA in agarose gel electrophoresis assays. These finding proves the ability of these benzo[a]phenoxazinium dyes to strongly interact with DNA.

  18. Evaluation of the probe dihydrocalcein acetoxymethylester as an indicator of reactive oxygen species formation and comparison with oxidative DNA base modification determined by modified alkaline elution technique.

    Science.gov (United States)

    Rohnstock, A; Lehmann, L

    2007-12-01

    Reactive oxygen species (ROS) play a predominant role in various diseases and the development of fast and easy methods for the quantification of intracellular ROS represents an important goal. Therefore, the aim of the present study was the evaluation of the fluorogenic probe dihydrocalcein acetoxymethylester (AM) for the detection of intracellular ROS. A flow cytometric method was developed using MCF-7 cells and the kinetics of ester hydrolysis and the cellular distribution and stability of calcein were characterized using calcein AM. Then, MCF-7 cells were challenged with model agents for the generation of singlet oxygen (illumination with visible light), peroxyl and hydroxyl radicals (tert-butylhydroperoxide, tBHP), superoxide anion radicals (potassium dioxide), and the intracellular formation of superoxide anion radicals by redox cycling (menadione) and the formation of calcein was compared with the induction of oxidative DNA base modifications assessed by modified alkaline elution technique. Every model agent significantly induced formamidopyrimidine-DNA glycosylase-sensitive sites (i.e. oxidative DNA base modifications) and most also induced DNA strand breaks. In contrast, exclusively tBHP and illumination with visible light induced the intracellular formation of calcein. In conclusion, though intracellular oxidation of dihydrocalcein represents a fast screening method, it detects a limited spectrum of ROS.

  19. Time-resolved luminescence biosensor for continuous activity detection of protein acetylation-related enzymes based on DNA-sensitized terbium(III) probes.

    Science.gov (United States)

    Han, Yitao; Li, Hao; Hu, Yufang; Li, Pei; Wang, Huixia; Nie, Zhou; Yao, Shouzhuo

    2015-09-15

    Protein acetylation of histone is an essential post-translational modification (PTM) mechanism in epigenetic gene regulation, and its status is reversibly controlled by histone acetyltransferases (HATs) and histone deacetylases (HDACs). Herein, we have developed a sensitive and label-free time-resolved luminescence (TRL) biosensor for continuous detection of enzymatic activity of HATs and HDACs, respectively, based on acetylation-mediated peptide/DNA interaction and Tb(3+)/DNA luminescent probes. Using guanine (G)-rich DNA-sensitized Tb(3+) luminescence as the output signal, the polycationic substrate peptides interact with DNA with high affinity and subsequently replace Tb(3+), eliminating the luminescent signal. HAT-catalyzed acetylation remarkably reduces the positive charge of the peptides and diminishes the peptide/DNA interaction, resulting in the signal on detection via recovery of DNA-sensitized Tb(3+) luminescence. With this TRL sensor, HAT (p300) can be sensitively detected with a wide linear range from 0.2 to 100 nM and a low detection limit of 0.05 nM. The proposed sensor was further used to continuously monitor the HAT activity in real time. Additionally, the TRL biosensor was successfully applied to evaluating HAT inhibition by two specific inhibitors, anacardic acid and C464, and satisfactory Z'-factors above 0.73 were obtained. Moreover, this sensor is feasible to continuously monitor the HDAC (Sirt1)-catalyzed deacetylation with a linear range from 0.5 to 500 nM and a detection limit of 0.5 nM. The proposed sensor is a convenient, sensitive, and mix-and-read assay, presenting a promising platform for protein acetylation-targeted epigenetic research and drug discovery.

  20. Label-free detection of specific DNA sequence-telomere using unmodified gold nanoparticles as colorimetric probes

    Science.gov (United States)

    Qi, Yingying; Li, Li; Li, Baoxin

    2009-09-01

    A simple and sensitive label-free colorimetric detection of telomere DNA has been developed. It was based on the color change of gold nanoparticles (AuNPs) due to DNA hybridization. UV-vis spectra and transmission electron microscopy (TEM) were used to investigate the change of AuNPs. Under the optimized conditions, the linear range for determination of telomere DNA was 5.7 × 10 -13 to 4.5 × 10 -6 mol/L. The detection limit (3 σ) of this method has decreased to pico-molar level.

  1. Genetic diversity of Helicobacter pylori indexed with respect to clinical symptomatology, using a 16S rRNA and a species-specific DNA probe.

    Science.gov (United States)

    Desai, M; Linton, D; Owen, R J; Cameron, H; Stanley, J

    1993-12-01

    DNA probes are described which identify group and fingerprint strains of the human gastric pathogen Helicobacter pylori, on the basis of well-defined band homologies. A 544 bp internal fragment of the 16S ribosomal RNA gene was generated by polymerase chain reaction (PCR) with primers derived from the Escherichia coli rRNA gene sequence. In genomic Southern blots this probe detected restriction site variation around these loci, generating simple but strain-specific molecular fingerprints. A small conserved chromosomal fragment of 1.2 kbp, Hps, species-specific for H. pylori, was obtained by cloning random HindIII fragments into pUC19. It was useful for dot-blot identification, and also separated isolates into one major and two minor groups. When results for these two probes were combined, a baseline characterization of genotype was obtained. A band-matching database of molecular fingerprints for the type strain and 63 clinical isolates of H. pylori from asymptomatic, ulcer and gastritis contexts is presented. No significant association between the genotypes at this level of definition and the associated clinical symptomatology of the isolates was detected.

  2. Hematopoietic Lineage Transcriptome Stability and Representation in PAXgeneTM Collected Peripheral Blood Utilising SPIA Single-Stranded cDNA Probes for Microarray

    Directory of Open Access Journals (Sweden)

    Laura Kennedy

    2008-01-01

    Full Text Available Peripheral blood as a surrogate tissue for transcriptome profiling holds great promise for the discovery of diagnostic and prognostic disease biomarkers, particularly when target tissues of disease are not readily available. To maximize the reliability of gene expression data generated from clinical blood samples, both the sample collection and the microarray probe generation methods should be optimized to provide stabilized, reproducible and representative gene expression profiles faithfully representing the transcriptional profiles of the constituent blood cell types present in the circulation. Given the increasing innovation in this field in recent years, we investigated a combination of methodological advances in both RNA stabilisation and microarray probe generation with the goal of achieving robust, reliable and representative transcriptional profiles from whole blood. To assess the whole blood profiles, the transcriptomes of purified blood cell types were measured and compared with the global transcriptomes measured in whole blood. The results demonstrate that a combination of PAXgeneTM RNA stabilising technology and single-stranded cDNA probe generation afforded by the NuGEN Ovation RNA amplification system V2TM enables an approach that yields faithful representation of specific hematopoietic cell lineage transcriptomes in whole blood without the necessity for prior sample fractionation, cell enrichment or globin reduction. Storage stability assessments of the PAXgeneTM blood samples also advocate a short, fixed room temperature storage time for all PAXgeneTM blood samples collected for the purposes of global transcriptional profiling in clinical studies.

  3. A Simple Method of Detecting Chlamydia Trachomatis Using Enzymatically Amplified DNA and Immobilized Probes on Microtiter Plate

    Institute of Scientific and Technical Information of China (English)

    王仁礼; 熊艳; 张龙兴; 蒋秀蓉; 张忠恕

    1998-01-01

    We have developed a simple and economical method for Chlamydia trachomatis detecting, called microtiter plate hybridization (PCR-MPtt) , which may replace standard PCR. This method is similar to that of an ELISA. Briefly, the PCR products labeled at the 5' termini with biotin were hybridized with probes immobilized on a microtiter well

  4. Quantitative super-resolution localization microscopy of DNA in situ using Vybrant® DyeCycle™ Violet fluorescent probe

    Directory of Open Access Journals (Sweden)

    Dominika Żurek-Biesiada

    2016-06-01

    Full Text Available Single Molecule Localization Microscopy (SMLM is a recently emerged optical imaging method that was shown to achieve a resolution in the order of tens of nanometers in intact cells. Novel high resolution imaging methods might be crucial for understanding of how the chromatin, a complex of DNA and proteins, is arranged in the eukaryotic cell nucleus. Such an approach utilizing switching of a fluorescent, DNA-binding dye Vybrant® DyeCycle™ Violet has been previously demonstrated by us (Żurek-Biesiada et al., 2015 [1]. Here we provide quantitative information on the influence of the chemical environment on the behavior of the dye, discuss the variability in the DNA-associated signal density, and demonstrate direct proof of enhanced structural resolution. Furthermore, we compare different visualization approaches. Finally, we describe various opportunities of multicolor DNA/SMLM imaging in eukaryotic cell nuclei.

  5. Quantitative super-resolution localization microscopy of DNA in situ using Vybrant® DyeCycle™ Violet fluorescent probe.

    Science.gov (United States)

    Żurek-Biesiada, Dominika; Szczurek, Aleksander T; Prakash, Kirti; Best, Gerrit; Mohana, Giriram K; Lee, Hyun-Keun; Roignant, Jean-Yves; Dobrucki, Jurek W; Cremer, Christoph; Birk, Udo

    2016-06-01

    Single Molecule Localization Microscopy (SMLM) is a recently emerged optical imaging method that was shown to achieve a resolution in the order of tens of nanometers in intact cells. Novel high resolution imaging methods might be crucial for understanding of how the chromatin, a complex of DNA and proteins, is arranged in the eukaryotic cell nucleus. Such an approach utilizing switching of a fluorescent, DNA-binding dye Vybrant® DyeCycle™ Violet has been previously demonstrated by us (Żurek-Biesiada et al., 2015) [1]. Here we provide quantitative information on the influence of the chemical environment on the behavior of the dye, discuss the variability in the DNA-associated signal density, and demonstrate direct proof of enhanced structural resolution. Furthermore, we compare different visualization approaches. Finally, we describe various opportunities of multicolor DNA/SMLM imaging in eukaryotic cell nuclei.

  6. Thermodynamic and structural study of pyrene-1-carboxaldehyde/DNA interactions by molecular spectroscopy: Probing intercalation and binding properties

    Science.gov (United States)

    Grueso, E.; Prado-Gotor, R.

    2010-08-01

    The binding of pyrene-1-carboxaldehyde (1-PyCHO) with ctDNA was investigated through absorption, intrinsic and induced circular dichroism, viscosity measurements and steady-state fluorescence. The binding and the number of monomer units of the polymer involved in the binding of one dye molecule (site size) have been quantified. The results indicated that the 1-PyCHO molecule binds to the ctDNA in an intercalative mode. The spectroscopic evidence of this intercalation process is also corroborated by the effect of urea, iodide-induced fluorescence quenching of pyrene-1-carboxaldehyde and competitive binding using a fluorescent intercalator, SYBR Green I (SG). The induced circular dichroism (ICD) spectra of pyrene-1-carboxaldehyde complexed with ctDNA show that pyrene-1-carboxaldehyde intercalates into ctDNA and that the intercalation orientation of pyrene to the DNA base-pairs long axis is heterogeneous. On the other hand, the intrinsic circular dichroism (CD) spectra show a stabilization of the right-handed B form of ctDNA, due to the intercalation process.

  7. Porphyrinic metal-organic framework as electrochemical probe for DNA sensing via triple-helix molecular switch.

    Science.gov (United States)

    Ling, Pinghua; Lei, Jianping; Ju, Huangxian

    2015-09-15

    An electrochemical DNA sensor was developed based on the electrocatalysis of porphyrinic metal-organic framework (MOF) and triple-helix molecular switch for signal transduction. The streptavidin functionalized zirconium-porphyrin MOF (PCN-222@SA) was prepared as signal nanoprobe via covalent method and demonstrated high electrocatalysis for O2 reduction. Due to the large steric effect, the designed nanoprobe was blocked for the interaction with the biotin labeled triple-helix immobilized on the surface of glassy carbon electrode. In the presence of target DNA, the assistant DNA in triple-helix will hybridize with target DNA, resulting in the disassembly of triple-helix molecular. Consequently, the end biotin away from the electrode was ''activated'' for easy access to the signal nanoprobe, PCN-222@SA, on the basis of biotin-streptavidin biorecognition. The introduction of signal nanoprobe to a sensor surface led to a significantly amplified electrocatalytic current towards oxygen reduction. Integrating with DNA recycling amplification of Exonuclease III, the sensitivity of the biosensor was improved significantly with detection limit of 0.29 fM. Moreover, the present method has been successfully applied to detect DNA in complex serum matrix. This porphyrinic MOF-based strategy has promising application in the determination of various analytes for signal transduction and has great potential in bioassays.

  8. Effect of fiberoptic bronchoscope compared with direct laryngoscope on hemodynamic responses to orotracheal intubation

    Institute of Scientific and Technical Information of China (English)

    ZHANG Guo-hua; XUE Fu-shan; LI Ping; SUN Hai-yan; LIU Kun-peng; XU Ya-chao; LIU Yi; SUN Hai-tao

    2007-01-01

    @@ Fiberoptic bronchoscope (FOB) is an important instrument for respiratory, disorder examination and difficult airway management.1 The fiberoptic intubation can avoid the mechanical stimulus to oropharyngolaryngeal structures thereby it is likely to attenuate hemodynamic responses during orotracheal intubation.

  9. Fiber-optically sensorized composite wing

    Science.gov (United States)

    Costa, Joannes M.; Black, Richard J.; Moslehi, Behzad; Oblea, Levy; Patel, Rona; Sotoudeh, Vahid; Abouzeida, Essam; Quinones, Vladimir; Gowayed, Yasser; Soobramaney, Paul; Flowers, George

    2014-04-01

    Electromagnetic interference (EMI) immune and light-weight, fiber-optic sensor based Structural Health Monitoring (SHM) will find increasing application in aerospace structures ranging from aircraft wings to jet engine vanes. Intelligent Fiber Optic Systems Corporation (IFOS) has been developing multi-functional fiber Bragg grating (FBG) sensor systems including parallel processing FBG interrogators combined with advanced signal processing for SHM, structural state sensing and load monitoring applications. This paper reports work with Auburn University on embedding and testing FBG sensor arrays in a quarter scale model of a T38 composite wing. The wing was designed and manufactured using fabric reinforced polymer matrix composites. FBG sensors were embedded under the top layer of the composite. Their positions were chosen based on strain maps determined by finite element analysis. Static and dynamic testing confirmed expected response from the FBGs. The demonstrated technology has the potential to be further developed into an autonomous onboard system to perform load monitoring, SHM and Non-Destructive Evaluation (NDE) of composite aerospace structures (wings and rotorcraft blades). This platform technology could also be applied to flight testing of morphing and aero-elastic control surfaces.

  10. Thermodynamic and structural study of phenanthroline derivative ruthenium complex/DNA interactions: probing partial intercalation and binding properties.

    Science.gov (United States)

    Grueso, E; López-Pérez, G; Castellano, M; Prado-Gotor, R

    2012-01-01

    The binding of [Ru(PDTA-H(2))(phen)]Cl (PDTA = propylene-1,2-diaminetetra-acetic acid; phen = 1,10 phenanthroline) with ctDNA (=calf thymus DNA) has been investigated through intrinsic and induced circular dichroism, UV-visible absorption and fluorescence spectroscopies, steady-state fluorescence, thermal denaturation technique, viscosity and electrochemical measurements. The latter indicate that the cathodic and anodic peak potentials of the ruthenium complex shift to more positive values on increasing the DNA concentration, this behavior being a direct consequence of the interaction of both the reduced and oxidized form with DNA binding. From spectrophotometric titration experiments, the equilibrium binding constant and the number of monomer units of the polymer involved in the binding of one ruthenium molecule (site size) have been quantified. The intrinsic circular dichroism (CD) spectra show an unwinding and a conformational change of the DNA helix upon interaction of the ruthenium complex. Quenching process, thermal denaturation experiments and induced circular dichroism (ICD) are consistent with a partial intercalative binding mode. Copyright © 2011 Elsevier Inc. All rights reserved.

  11. Fiber-optic ultrasonic hydrophone using short Fabry-Perot cavity with multilayer reflectors deposited on small stub.

    Science.gov (United States)

    Kim, Kyung-Su; Mizuno, Yosuke; Nakamura, Kentaro

    2014-04-01

    A fiber-optic probe with dielectric multilayer films deposited on a small stub is studied for mega-hertz ultrasonic-wave detection in water. The small stub with a short Fabry-Perot cavity and distributed reflectors is attached on the fiber end. The structure is mechanically strong and withstands intense ultrasonic pressure. Ultrasonic waves at 1.56MHz are successfully detected in water with a good signal-to-noise ratio. The working principle and the characteristics are studied by comparing the ultrasonic sensitivity with that of a conventional piezoelectric hydrophone. The distance response and directional response are also investigated.

  12. A Noninvasive Miniaturized-Wireless Laser-Doppler Fiber-Optic Sensor for Understanding Distal Fingertip Injuries in Astronauts

    Science.gov (United States)

    Ansari, Rafat R.; Jones, Jeffrey A.; Pollonini, Luca; Rodriquez, Mikael; Opperman, Roedolph; Hochstein, Jason

    2009-01-01

    During extra-vehicular activities (EVAs) or spacewalks astronauts over use their fingertips under pressure inside the confined spaces of gloves/space suits. The repetitive hand motion is a probable cause for discomfort and injuries to the fingertips. We describe a new wireless fiber-optic probe that can be integrated inside the astronaut glove for noninvasive blood perfusion measurements in distal fingertips. In this preliminary study, we present blood perfusion measurements while performing hand-grip exercises simulating the use of space tools.

  13. Bacteria capable of degrading anthracene, phenanthrene, and fluoranthene as revealed by DNA based stable-isotope probing in a forest soil

    Energy Technology Data Exchange (ETDEWEB)

    Song, Mengke [Guangzhou Institute of Geochemistry, Chinese Academy of Sciences, Guangzhou 510640 (China); Jiang, Longfei [College of Life Sciences, Nanjing Agricultural University, Nanjing 210095 (China); Zhang, Dayi [Lancaster Environment Centre, Lancaster University, Lancaster LA1 4YQ (United Kingdom); Luo, Chunling, E-mail: clluo@gig.ac.cn [Guangzhou Institute of Geochemistry, Chinese Academy of Sciences, Guangzhou 510640 (China); Wang, Yan [Key Laboratory of Industrial Ecology and Environmental Engineering (MOE), School of Environmental Science and Technology, Dalian University of Technology, Dalian 116024 (China); Yu, Zhiqiang [Guangzhou Institute of Geochemistry, Chinese Academy of Sciences, Guangzhou 510640 (China); Yin, Hua [College of Environment and Energy, South China University of Technology, Guangzhou 510006 (China); Zhang, Gan [Guangzhou Institute of Geochemistry, Chinese Academy of Sciences, Guangzhou 510640 (China)

    2016-05-05

    Highlights: • Investigate PAHs degraders in forest carbon-rich soils via DNA-SIP. • Rhodanobacter is identified to metabolite anthracene for the first time. • The first fluoranthene degrader belongs to Acidobacteria. • Different functions of PAHs degraders in forest soils from contaminated soils. - Abstract: Information on microorganisms possessing the ability to metabolize different polycyclic aromatic hydrocarbons (PAHs) in complex environments helps in understanding PAHs behavior in natural environment and developing bioremediation strategies. In the present study, stable-isotope probing (SIP) was applied to investigate degraders of PAHs in a forest soil with the addition of individually {sup 13}C-labeled phenanthrene, anthracene, and fluoranthene. Three distinct phylotypes were identified as the active phenanthrene-, anthracene- and fluoranthene-degrading bacteria. The putative phenanthrene degraders were classified as belonging to the genus Sphingomona. For anthracene, bacteria of the genus Rhodanobacter were the putative degraders, and in the microcosm amended with fluoranthene, the putative degraders were identified as belonging to the phylum Acidobacteria. Our results from DNA-SIP are the first to directly link Rhodanobacter- and Acidobacteria-related bacteria with anthracene and fluoranthene degradation, respectively. The results also illustrate the specificity and diversity of three- and four-ring PAHs degraders in forest soil, contributes to our understanding on natural PAHs biodegradation processes, and also proves the feasibility and practicality of DNA-based SIP for linking functions with identity especially uncultured microorganisms in complex microbial biota.

  14. Restriction fragment length polymorphism and multiple copies of DNA sequences homologous with probes for P-fimbriae and hemolysin genes among uropathogenic Escherichia coli.

    Science.gov (United States)

    Hull, S I; Bieler, S; Hull, R A

    1988-03-01

    Hemolysin and P-fimbriae are two virulence traits frequently found together in uropathogenic Escherichia coli. Previous studies have discovered evidence both for linkage between the genes for these traits and for their duplication in the chromosomes of a limited number of strains. To test whether these observations are characteristic of uropathogenic Escherichia coli, the method of DNA hybridization to DNA restriction fragments separated by electrophoresis and transferred to nylon was used to determine copy number of genes for P-fimbriae (pap) among 51 E. coli strains isolated from symptomatic urinary tract infections. Twenty percent of the strains had more than one copy of pap homologous sequences. Fifteen strains, each representing a unique clone, were examined for the presence of sequences homologous with cloned hemolysin genes (hly). Samples of DNA from 14 of the 15 strains hybridized with hly probes. In eight strains the number of copies of pap equalled the number of copies of hly, including one strain with two apparent copies of each. Five strains appeared to have one more copy of pap than of hly, and one strain had an extra copy of hly.

  15. Screening cDNA Libraries Using Partial Probes to Isolate Full-Length cDNAs from Vascular Cells.

    Science.gov (United States)

    Csortos, C; Lazar, V; Garcia, J G

    1999-01-01

    The purpose of screening cDNA libraries is to isolate a particular cDNA clone encoding a mRNA and by implication, a protein, of interest. The screening is based on identification of the desired clone among a large number of recombinant clones within the library selected (1,2). As an example of both the utility and power of library screening, we will relate our own library screening efforts utilized to isolate the nonmuscle high molecular weight myosin light chain kinase isoform from a human umbilical vein endothelial cell cDNA library (3). This unique nonmuscle myosin light chain kinase isoform phosphorylates myosin light chains, thereby playing an essential role in agonist-mediated endothelial cell contraction, paracellular gap formation and increased vascular permeability. We are hopeful that this step-by-step approach will help the reader to understand the discussed methods.

  16. Incorporation of nucleoside probes opposite O⁶-methylguanine by Sulfolobus solfataricus DNA polymerase Dpo4: importance of hydrogen bonding.

    Science.gov (United States)

    Stornetta, Alessia; Angelov, Todor; Guengerich, F Peter; Sturla, Shana J

    2013-09-02

    O⁶-Methylguanine (O⁶-MeG) is a mutagenic DNA lesion, arising from the action of methylating agents on guanine (G) in DNA. Dpo4, an archaeal low-fidelity Y-family DNA polymerase involved in translesion DNA synthesis (TLS), is a model for studying how human Y-family polymerases bypass DNA adducts. Previous work showed that Dpo4-mediated dTTP incorporation is favored opposite O⁶-MeG rather than opposite G. However, factors influencing the preference of Dpo4 to incorporate dTTP opposite O⁶-MeG are not fully defined. In this study, we investigated the influence of structural features of incoming dNTPs on their enzymatic incorporation opposite O⁶-MeG in a DNA template. To this end, we utilized a new fluorescence-based primer extension assay to evaluate the incorporation efficiency of a panel of synthetic dNTPs opposite G or O⁶-MeG by Dpo4. In single-dNTP primer extension studies, the synthetic dNTPs were preferentially incorporated opposite G, relative to O⁶-MeG. Moreover, pyrimidine-based dNTPs were generally better incorporated than purine-based syn-conformation dNTPs. The results suggest that hydrophobicity of the incoming dNTP appears to have little influence on the process of nucleotide selection by Dpo4, with hydrogen bonding capacity being a major influence. Additionally, modifications at the C2-position of dCTP increase the selectivity for incorporation opposite O⁶-MeG without a significant loss of efficiency.

  17. Molecular Mechanism of Dioxin Action: Molecular Cloning of the Ah Receptor Using a DNA Recognition Site Probe

    Science.gov (United States)

    1992-01-13

    DNA with high affinity (Whitlock and Galeazzi , 1984; Henry et al., 1989; Denison and Yao, 1991). Biochemical and genetic studies (Denison et al., 1988a...Pharmacol. Toxicol. 30: 251-277. Whitlock, J. P., Jr. (1987) Pharmacol. Rev. 39: 147-161. Whitlock, J. P., Jr. and Galeazzi , D. R. (1984) J. Biol

  18. Fiber-optic nonlinear endomicroscopy with focus scanning by using shape memory alloy actuation

    Science.gov (United States)

    Wu, Yicong; Zhang, Yuying; Xi, Jiefeng; Li, Ming-Jun; Li, Xingde

    2010-01-01

    A miniature fiber optic endomicroscope with built-in dynamic focus scanning capability is developed for the first time for 3-D two-photon fluorescence (TPF) imaging of biological samples. Fast 2-D lateral beam scanning is realized by resonantly vibrating a double-clad fiber cantilever with a tubular piezoactuator. Slow axial scanning is achieved by moving the distal end of the imaging probe with an extremely compact electrically driven shape memory alloy (SMA). The 10-mm-long SMA allows 150-μm contractions with a driving voltage varying only from 50 to 100 mV. The response of the SMA contraction with the applied voltage is nonlinear, but repeatable and can be accurately calibrated. Depth-resolved imaging of acriflavine-stained biological tissues and unstained white paper with the endomicroscope is performed, and the results demonstrate the feasibility of 3-D nonlinear optical imaging with the SMA-based scanning fiber-optic endomicroscope. PMID:21198147

  19. Cloning and characterization of cDNA probes for the analysis of metallothionein gene expression in the Mediterranean bivalves: Ruditapes decussatus and Cerastoderma glaucum.

    Science.gov (United States)

    Ladhar-Chaabouni, Rim; Mokdad-Gargouri, Raja; Denis, Françoise; Hamza-Chaffai, Amel

    2009-05-01

    cDNA probes have been developed for subsequent use in monitoring the cadmium exposure of the clam Ruditapes decussatus and the cockle Cerastoderma glaucum using metallothionein (MT) gene expression in different tissues of these species. Two partial MT cDNAs were isolated from Ruditapes decussatus and Cerastoderma glaucum. The identification of the nucleotide sequences showed that the cDNAs consist of 480 bp coding 72 amino acid proteins containing 21 cysteine residues organized in Cys-X-Cys motifs as classically described for MTs. The induction of MT gene expression in CdCl(2) treated bivalves was confirmed by dot blot analysis and suggests a potential specific tissue expression rate.

  20. Fiber-Optic Terahertz Data-Communication Networks

    Science.gov (United States)

    Chua, Peter L.; Lambert, James L.; Morookian, John M.; Bergman, Larry A.

    1994-01-01

    Network protocols implemented in optical domain. Fiber-optic data-communication networks utilize fully available bandwidth of single-mode optical fibers. Two key features of method: use of subpicosecond laser pulses as carrier signals and spectral phase modulation of pulses for optical implementation of code-division multiple access as multiplexing network protocol. Local-area network designed according to concept offers full crossbar functionality, security of data in transit through network, and capacity about 100 times that of typical fiber-optic local-area network in current use.

  1. Dignostic Yield of Fiberoptic Bronchoscopy in a Teaching Hospital

    Directory of Open Access Journals (Sweden)

    Rajinder Singh

    2008-10-01

    Full Text Available Fiberoptic bronchoscopy is minimally invasive procedure which can be performed on outpatient basis.Thestudy is a reterospective review of the data at a tertiary center and compares the diganostic yield of thepatients (n=720, who underwent FB at our pulmonary unit with the data from international centers. Thediagnostic yield of the FB was high(70% with good selection of the patients and growth was the mostcommon finding followed by infections.FB was normal in 218(30% patients. Flexible fiberoptic bronchoscopyis a useful diagnostic tool with a low rate of complications. The diagnostic yield in our institution is alsmostsimilar to that reported in other series.

  2. Specific and Sensitive Isothermal Electrochemical Biosensor for Plant Pathogen DNA Detection with Colloidal Gold Nanoparticles as Probes

    Science.gov (United States)

    Lau, Han Yih; Wu, Haoqi; Wee, Eugene J. H.; Trau, Matt; Wang, Yuling; Botella, Jose R.

    2017-01-01

    Developing quick and sensitive molecular diagnostics for plant pathogen detection is challenging. Herein, a nanoparticle based electrochemical biosensor was developed for rapid and sensitive detection of plant pathogen DNA on disposable screen-printed carbon electrodes. This 60 min assay relied on the rapid isothermal amplification of target pathogen DNA sequences by recombinase polymerase amplification (RPA) followed by gold nanoparticle-based electrochemical assessment with differential pulse voltammetry (DPV). Our method was 10,000 times more sensitive than conventional polymerase chain reaction (PCR)/gel electrophoresis and could readily identify P. syringae infected plant samples even before the disease symptoms were visible. On the basis of the speed, sensitivity, simplicity and portability of the approach, we believe the method has potential as a rapid disease management solution for applications in agriculture diagnostics.

  3. Suppression subtractive hybridization: a method for generating differentially regulated or tissue-specific cDNA probes and libraries.

    OpenAIRE

    1996-01-01

    A new and highly effective method, termed suppression subtractive hybridization (SSH), has been developed for the generation of subtracted cDNA libraries. It is based primarily on a recently described technique called suppression PCR and combines normalization and subtraction in a single procedure. The normalization step equalizes the abundance of cDNAs within the target population and the subtraction step excludes the common sequences between the target and driver populations. In a model sys...

  4. Probe into the Construction of Criminal DNA Database%犯罪DNA数据库建设的探讨

    Institute of Scientific and Technical Information of China (English)

    2013-01-01

      自1995年世界上第一个国家DNA数据库在英国建立后,全世界的DNA数据库建设已有了很大的发展,犯罪DNA数据库在侦查破案等工作中的作用更加突出。我国的犯罪DNA数据库是为了帮助公安机关高效执法而建立的,数据库所起的锁定刑事案件犯罪嫌疑人的作用往往是其他手段与方法很难达到的。在司法鉴定领域,遗传基因分析技术也广泛地应用于个体识别和鉴定之中。建立犯罪DNA数据库已经作为刑事案件侦破的有效手段,其技术已走向成熟。%Since the first national DNA database in the world was established in UK in 1995, the world has witnessed great progress in the construction of DNA databases in various countries, which has an outstanding effect on criminal investigation. The DNA databases in our country are constructed mainly to help police agencies in their law enforcement. Unlike other means of investigation, the DNA database has a unique function of identifying criminal suspects. In the forensic science, the genetic analytic technique is now widely used in paternity test and personal identification. Its establishment is an important means of criminal investigation and its techniques are increasingly perfect.

  5. Probing the functional impact of sequence variation on p53-DNA interactions using a novel microsphere assay for protein-DNA binding with human cell extracts.

    Directory of Open Access Journals (Sweden)

    Maher A Noureddine

    2009-05-01

    Full Text Available The p53 tumor suppressor regulates its target genes through sequence-specific binding to DNA response elements (REs. Although numerous p53 REs are established, the thousands more identified by bioinformatics are not easily subjected to comparative functional evaluation. To examine the relationship between RE sequence variation -- including polymorphisms -- and p53 binding, we have developed a multiplex format microsphere assay of protein-DNA binding (MAPD for p53 in nuclear extracts. Using MAPD we measured sequence-specific p53 binding of doxorubicin-activated or transiently expressed p53 to REs from established p53 target genes and p53 consensus REs. To assess the sensitivity and scalability of the assay, we tested 16 variants of the p21 target sequence and a 62-multiplex set of single nucleotide (nt variants of the p53 consensus sequence and found many changes in p53 binding that are not captured by current computational binding models. A group of eight single nucleotide polymorphisms (SNPs was examined and binding profiles closely matched transactivation capability tested in luciferase constructs. The in vitro binding characteristics of p53 in nuclear extracts recapitulated the cellular in vivo transactivation capabilities for eight well-established human REs measured by luciferase assay. Using a set of 26 bona fide REs, we observed distinct binding patterns characteristic of transiently expressed wild type and mutant p53s. This microsphere assay system utilizes biologically meaningful cell extracts in a multiplexed, quantitative, in vitro format that provides a powerful experimental tool for elucidating the functional impact of sequence polymorphism and protein variation on protein/DNA binding in transcriptional networks.

  6. THE CLONING OF HRNT-1 USING A COMBINATION OF cDNA LIBRARY SCREENING WITH BIOTIN-LABELED PROBE AND RAPID AMPLIFICATION OF cDNA ENDS

    Institute of Scientific and Technical Information of China (English)

    ZHANG; Kai-tai

    2001-01-01

    [1]Tom S, Andrew PR. Human Molecular Genetics [M]. John Wiley & Sons, Inc. United States of America 1996; 335.[2]Zhao Yong-liang, Jin Cui-zhen, Wu De-chang et al. Neoplastic transformation and cytogenetic changes of rat tracheal epithelial cells induced by a-particles irradiation [J]. Chin Med Sci J 1997; 12:202.[3]Frohman MA. Rapid amplification of complementary DNA ends for generation of full-length complementary DNAs: thermal RACE [J]. Methods Enzymol 1993; 218:340.[4]Frederick A, Roger B. Current Protocols in Molecular Biology [M]. John Wiley & Sons, Inc. United States of America 1998; 2.1.1.[5]Roux KH. Optimization and troubleshooting in PCR [J]. PCR Methods Appl 1995; 4:5158.[6]Sambrook, J, Fritsch EF, Maniatis T. Molecular Cloning: A Laboratory Manual [M]. 2nd Ed. New York: Cold Spring Harbor Laboratory, Cold Spring Harbor, 1989; 54.[7]Zhang Y, Frohman MA. Using rapid amplification of cDNA ends (RACE) to obtain full-length cDNAs [J]. Methods Mol Biol 1997; 69:61.[8]Frohman MA. Rapid amplification of complementary DNA ends for generation of full-length complementary DNAs: thermal RACE [J]. Methods Enzymol 1993; 218:340.[9]Iqbal S, Robinson J, Deere D, et al. Efficiency of the polymerase chain reaction amplification of the uid gene for detection of Escherichia coli in contaminated water [J]. Lett Appl Microbiol 1997; 24:498.[10]Schunck B, Kraft W, Truyen U. A simple touch-down polymerase chain reaction for the detection of canine parvovirus and feline panleukopenia virus in feces [J]. J Virol Methods 1995; 55:427.

  7. Combination of ICP-MS, capillary electrophoresis, and their hyphenation for probing Ru(III) metallodrug-DNA interactions.

    Science.gov (United States)

    Foteeva, Lidia S; Matczuk, Magdalena; Pawlak, Katarzyna; Aleksenko, Svetlana S; Nosenko, Sergey V; Karandashev, Vasily K; Jarosz, Maciej; Timerbaev, Andrei R

    2017-03-01

    Determination of the DNA-binding reactivity and affinity is an important part of a successful program for the selection of metallodrug candidates. For such assaying, a range of complementary analytical techniques was proposed and tested here using one of few anticancer metal-based drugs that are currently in clinical trials, indazolium trans-[tetrachloridobis(1H-indazole)ruthenate(III), and a DNA oligonucleotide. A high reactivity of the Ru drug was confirmed in affinity capillary electrophoresis (CE) mode, where adduct formation takes place in situ (i.e., in the capillary filled with an oligonucleotide-containing electrolyte). To further characterize the binding kinetics, a drug-oligonucleotide mixture was incubated for a different period of time, followed by ultrafiltration separation into two different in molecular weight fractions (>3 and ICP-MS), revealing that at least two DNA adducts exist at equilibrium conditions. Using standalone ICP-MS, dominant equilibrium amount of the bound ruthenium was found to occur in a fraction of 5-10 kDa, which includes the oligonucleotide (ca. 6 kDa). Importantly, in all three assays, the drug was used for the first time in in-vitro studies, not in the intact form but as its active species released from the transferrin adduct at simulated cancer cytosolic conditions. This circumstance makes the established analytical platform promising to provide a detailed view on metallodrug targeting, including other possible biomolecules and ex vivo samples.

  8. 功能化CdS∶Mn荧光探针测定hs-DNA%Detection of hs-DNA by functinal CdS∶Mn fluorescent probe

    Institute of Scientific and Technical Information of China (English)

    张艺; 李紫薇; 何晓燕; 王兴磊

    2015-01-01

    在油胺溶剂中,利用溶剂热法于160℃加热反应12 h,合成了CdS∶Mn量子点,并用巯基乙酸对合成的量子点进行了包裹.借助X射线衍射分析、傅里叶红外光谱、荧光谱图等测试方法对材料进行表征,数据表明,合成的CdS∶Mn量子点JCPDS#为80-0019,在CdS∶Mn量子点上成功修饰上了羧基(-COOH),CdS∶Mn晶体修饰前后主要发射峰从526 nm蓝移至488 nm.以COOH-CdS∶Mn功能化材料为荧光探针,基于DNA对材料的荧光猝灭,实现了对DNA的定量测定.实验在pH=6.6条件下,功能化荧光材料的荧光强度和鲱鱼精DNA(hs-DNA)的质量浓度呈线性关系,线性方程为△F =78.69-0.179c,检出限为0.053 mg/L,相关系数R=0.998 8,RSD =0.27%.

  9. Prevalence of transposons encoding kanamycin, ampicillin and trimethoprim resistance in isolates from urinary tract infections detected using DNA probes.

    Science.gov (United States)

    Chang, S F; Chang, L L; Chow, T Y; Wu, W J; Chang, J C

    1992-03-01

    Drug resistant Gram-negative bacteria causing urinary tract infections were collected. Kanamycin, ampicillin or trimethoprim-resistant strains were analyzed separately for the presence of Tn5, Tn3, or Tn7 by colony hybridization. Of these isolates, kanamycin-resistant transposons were present in 38.2% of 60 kanamycin-resistant isolates. A 3.3 kb fragment containing SacI-BamHI transposase of Tn3 and 42.6% showed a positive reaction in 129 ampicillin-resistant clinical isolates. Among the 75 trimethoprim-resistant isolates studied, 52% were shown to contain Tn7 when probed with a 1 kb BamHI fragment of Tn7. Results from Southern hybridizations demonstrated that these antibiotic resistant genes had been born on plasmids in some clinical isolates.

  10. In-plane ultrasonic needle tracking using a fiber-optic hydrophone

    Energy Technology Data Exchange (ETDEWEB)

    Xia, Wenfeng, E-mail: wenfeng.xia@ucl.ac.uk; Desjardins, Adrien E. [Department of Medical Physics and Biomedical Engineering, University College London, Gower Street, London WC1E 6BT (United Kingdom); Mari, Jean Martial [Department of Medical Physics and Biomedical Engineering, University College London, Gower Street, London WC1E 6BT, United Kingdom and GePaSud, University of French Polynesia, Faa’a 98702, French Polynesia (France); West, Simeon J. [Department of Anaesthesia, University College Hospital, Main Theatres, Maple Bridge Link Corridor, Podium 3, 235 Euston Road, London NW1 2BU (United Kingdom); Ginsberg, Yuval; David, Anna L. [Institute for Women’s Health, University College London, 86-96 Chenies Mews, London WC1E 6HX (United Kingdom); Ourselin, Sebastien [Center for Medical Imaging Computing, University College London, Gower Street, London WC1E 6BT (United Kingdom)

    2015-10-15

    Purpose: Accurate and efficient guidance of needles to procedural targets is critically important during percutaneous interventional procedures. Ultrasound imaging is widely used for real-time image guidance in a variety of clinical contexts, but with this modality, uncertainties about the location of the needle tip within the image plane lead to significant complications. Whilst several methods have been proposed to improve the visibility of the needle, achieving accuracy and compatibility with current clinical practice is an ongoing challenge. In this paper, the authors present a method for directly visualizing the needle tip using an integrated fiber-optic ultrasound receiver in conjunction with the imaging probe used to acquire B-mode ultrasound images. Methods: Needle visualization and ultrasound imaging were performed with a clinical ultrasound imaging system. A miniature fiber-optic ultrasound hydrophone was integrated into a 20 gauge injection needle tip to receive transmissions from individual transducer elements of the ultrasound imaging probe. The received signals were reconstructed to create an image of the needle tip. Ultrasound B-mode imaging was interleaved with needle tip imaging. A first set of measurements was acquired in water and tissue ex vivo with a wide range of insertion angles (15°–68°) to study the accuracy and sensitivity of the tracking method. A second set was acquired in an in vivo swine model, with needle insertions to the brachial plexus. A third set was acquired in an in vivo ovine model for fetal interventions, with insertions to different locations within the uterine cavity. Two linear ultrasound imaging probes were used: a 14–5 MHz probe for the first and second sets, and a 9–4 MHz probe for the third. Results: During insertions in tissue ex vivo and in vivo, the imaged needle tip had submillimeter axial and lateral dimensions. The signal-to-noise (SNR) of the needle tip was found to depend on the insertion angle. With

  11. Bacteria capable of degrading anthracene, phenanthrene, and fluoranthene as revealed by DNA based stable-isotope probing in a forest soil.

    Science.gov (United States)

    Song, Mengke; Jiang, Longfei; Zhang, Dayi; Luo, Chunling; Wang, Yan; Yu, Zhiqiang; Yin, Hua; Zhang, Gan

    2016-05-05

    Information on microorganisms possessing the ability to metabolize different polycyclic aromatic hydrocarbons (PAHs) in complex environments helps in understanding PAHs behavior in natural environment and developing bioremediation strategies. In the present study, stable-isotope probing (SIP) was applied to investigate degraders of PAHs in a forest soil with the addition of individually (13)C-labeled phenanthrene, anthracene, and fluoranthene. Three distinct phylotypes were identified as the active phenanthrene-, anthracene- and fluoranthene-degrading bacteria. The putative phenanthrene degraders were classified as belonging to the genus Sphingomona. For anthracene, bacteria of the genus Rhodanobacter were the putative degraders, and in the microcosm amended with fluoranthene, the putative degraders were identified as belonging to the phylum Acidobacteria. Our results from DNA-SIP are the first to directly link Rhodanobacter- and Acidobacteria-related bacteria with anthracene and fluoranthene degradation, respectively. The results also illustrate the specificity and diversity of three- and four-ring PAHs degraders in forest soil, contributes to our understanding on natural PAHs biodegradation processes, and also proves the feasibility and practicality of DNA-based SIP for linking functions with identity especially uncultured microorganisms in complex microbial biota.

  12. A Non-Invasive Multichannel Hybrid Fiber-Optic Sensor System for Vital Sign Monitoring

    Directory of Open Access Journals (Sweden)

    Marcel Fajkus

    2017-01-01

    Full Text Available In this article, we briefly describe the design, construction, and functional verification of a hybrid multichannel fiber-optic sensor system for basic vital sign monitoring. This sensor uses a novel non-invasive measurement probe based on the fiber Bragg grating (FBG. The probe is composed of two FBGs encapsulated inside a polydimethylsiloxane polymer (PDMS. The PDMS is non-reactive to human skin and resistant to electromagnetic waves, UV absorption, and radiation. We emphasize the construction of the probe to be specifically used for basic vital sign monitoring such as body temperature, respiratory rate and heart rate. The proposed sensor system can continuously process incoming signals from up to 128 individuals. We first present the overall design of this novel multichannel sensor and then elaborate on how it has the potential to simplify vital sign monitoring and consequently improve the comfort level of patients in long-term health care facilities, hospitals and clinics. The reference ECG signal was acquired with the use of standard gel electrodes fixed to the monitored person's chest using a real-time monitoring system for ECG signals with virtual instrumentation. The outcomes of these experiments have unambiguously proved the functionality of the sensor system and will be used to inform our future research in this fast developing and emerging field.

  13. A Non-Invasive Multichannel Hybrid Fiber-Optic Sensor System for Vital Sign Monitoring

    Science.gov (United States)

    Fajkus, Marcel; Nedoma, Jan; Martinek, Radek; Vasinek, Vladimir; Nazeran, Homer; Siska, Petr

    2017-01-01

    In this article, we briefly describe the design, construction, and functional verification of a hybrid multichannel fiber-optic sensor system for basic vital sign monitoring. This sensor uses a novel non-invasive measurement probe based on the fiber Bragg grating (FBG). The probe is composed of two FBGs encapsulated inside a polydimethylsiloxane polymer (PDMS). The PDMS is non-reactive to human skin and resistant to electromagnetic waves, UV absorption, and radiation. We emphasize the construction of the probe to be specifically used for basic vital sign monitoring such as body temperature, respiratory rate and heart rate. The proposed sensor system can continuously process incoming signals from up to 128 individuals. We first present the overall design of this novel multichannel sensor and then elaborate on how it has the potential to simplify vital sign monitoring and consequently improve the comfort level of patients in long-term health care facilities, hospitals and clinics. The reference ECG signal was acquired with the use of standard gel electrodes fixed to the monitored person’s chest using a real-time monitoring system for ECG signals with virtual instrumentation. The outcomes of these experiments have unambiguously proved the functionality of the sensor system and will be used to inform our future research in this fast developing and emerging field. PMID:28075341

  14. Development of a Single Stranded DNA Aptamer as a Molecular Probe for LNCap Cells Using Cell-SELEX.

    Science.gov (United States)

    Almasi, Faezeh; Mousavi Gargari, Seyed Latif; Bitaraf, Fatemeh; Rasoulinejad, Samaneh

    2016-01-01

    Nowadays, highly specific aptamers generated by cell SELEX technology (systematic evolution of ligands by exponential enrichment) are being applied for early detection of cancer cells. Prostate Specific Membrane Antigen (PSMA), over expressed in prostate cancer, is a highly specific marker and therefore can be used for diagnosis of the prostate cancer cells. The aim of the present study was to select single-stranded DNA aptamers against LNCap cells highly expressing PSMA, using cell-SELEX method which can be used as a diagnostic tool for the detection of prostate cancer cells. After 10 rounds of cell-SELEX, DNA aptamers were isolated against PSMA using LNCaP cells as a target and PC-3 cell lines for counter SELEX. Five DNA aptamers with more than 70% affinity were selected up on flow cytometry analysis of positive clones. Dissociation constants of two selected sequences (A12-B1) were estimated in the range of 33.78±3.77 and 57.49±2.214 pmol, respectively. Conserved secondary structures of A12 and B1 sequences suggest the necessity of these structures for binding with high affinity to native PSMA. Comparison of the secondary structures of our isolated aptamers and aptamer A10 obtained by protein SELEX showed similar stem-loop structures which could be responsible for the recognition of PSMA on LNCap cell surface. Our results indicated that selected aptamers may turn out to be ideal candidates for the development of a detection tool and also can be used in targeted drug delivery for future smart drugs.

  15. Isolation of a human anti-haemophilic factor IX cDNA clone using a unique 52-base synthetic oligonucleotide probe deduced from the amino acid sequence of bovine factor IX.

    Science.gov (United States)

    Jaye, M; de la Salle, H; Schamber, F; Balland, A; Kohli, V; Findeli, A; Tolstoshev, P; Lecocq, J P

    1983-04-25

    A unique 52mer oligonucleotide deduced from the amino acid sequence of bovine Factor IX was synthesized and used as a probe to screen a human liver cDNA bank. The Factor IX clone isolated shows 5 differences in nucleotide and deduced amino acid sequence as compared to a previously isolated clone. In addition, precisely one codon has been deleted.Images

  16. “大范围抽血采验DNA”现象研究%Probe on "DNA Dragnet"

    Institute of Scientific and Technical Information of China (English)

    王志刚

    2012-01-01

    随着DNA技术在刑事司法实践中的应用,"大范围抽血采验DNA"这种侦查方式被侦查机关广泛使用,其对刑事案件的侦破起到了一定积极作用,但也存在较大缺陷。根据对"大范围抽血采验DNA"的产生发展及其实际应用状况等方面的分析,这一侦查方式不仅侵害公民合法权益,而且成本高昂、成功机率不高。应规范侦查机关的侦查权,完善刑事诉讼法。%Along with DNA technology application in criminal judicial practice, this investigation measure was widely used and played a certain positive role in investigation, however, there are also obvious flaws in it. Based on the analysis from the aspects of generation, development and practical application of "DNA Drag- nets", we can find that this measure not only infringes citizens' legitimate rights, but also costs much and its success rate is not high. The power of the investigation organs should be regulated and Chinese criminal proce- dure law should be perfected.

  17. Kansas Communication and Instruction System through Fiber-Optic Transmission.

    Science.gov (United States)

    Kansas State Dept. of Education, Topeka.

    Schools and communities will restructure as they move into the next decade. The success of this restructuring will be dependent upon access to and sharing of quality teaching and information through an expanded communication system. One of the major two-way interactive technologies is the fiber-optic cable: a delivery system that will provide…

  18. Malposition of the epiglottis associated with fiberoptic intubation.

    Science.gov (United States)

    Takenaka, Ichiro; Aoyama, Kazuyoshi; Abe, Yumiko; Iwagaki, Tamao; Takenaka, Yukari; Kadoya, Tatsuo

    2009-02-01

    A case in which the epiglottis was tucked into the laryngeal inlet by advancement of an endotracheal tube (ETT) during fiberoptic intubation, is presented. In this case, pulling the fibroscope, which was advanced under the displaced epiglottis, was effective for restoration.

  19. Distributed Fiber-Optic Sensors for Vibration Detection.

    Science.gov (United States)

    Liu, Xin; Jin, Baoquan; Bai, Qing; Wang, Yu; Wang, Dong; Wang, Yuncai

    2016-07-26

    Distributed fiber-optic vibration sensors receive extensive investigation and play a significant role in the sensor panorama. Optical parameters such as light intensity, phase, polarization state, or light frequency will change when external vibration is applied on the sensing fiber. In this paper, various technologies of distributed fiber-optic vibration sensing are reviewed, from interferometric sensing technology, such as Sagnac, Mach-Zehnder, and Michelson, to backscattering-based sensing technology, such as phase-sensitive optical time domain reflectometer, polarization-optical time domain reflectometer, optical frequency domain reflectometer, as well as some combinations of interferometric and backscattering-based techniques. Their operation principles are presented and recent research efforts are also included. Finally, the applications of distributed fiber-optic vibration sensors are summarized, which mainly include structural health monitoring and perimeter security, etc. Overall, distributed fiber-optic vibration sensors possess the advantages of large-scale monitoring, good concealment, excellent flexibility, and immunity to electromagnetic interference, and thus show considerable potential for a variety of practical applications.

  20. DNA

    Science.gov (United States)

    Stent, Gunther S.

    1970-01-01

    This history for molecular genetics and its explanation of DNA begins with an analysis of the Golden Jubilee essay papers, 1955. The paper ends stating that the higher nervous system is the one major frontier of biological inquiry which still offers some romance of research. (Author/VW)

  1. Temperature-independent refractometer based on fiber-optic Fabry-Perot interferometer

    Science.gov (United States)

    Li, Jiacheng; Qiao, Xueguang; Wang, Ruohui; Rong, Qiangzhou; Bao, Weijia; Shao, Zhihua; Yang, Tingting

    2016-04-01

    A miniature fiber-optic refractometer based on Fabry-Perot interferometer (FPI) has been proposed and experimentally demonstrated. The sensing head consists of a short section of photonics crystal fiber (PCF) spliced to a single mode fiber (SMF), in which the end-face of the PCF is etched to remove holey structure with hydrofluoric (HF) acid. A Fabry-Perot interference spectrum is achieved based on the reflections from the fusion splicing interface and the end-face of the core of PCF. The interference fringe is sensitive to the external refractive index (RI) with an intensity-referenced sensitivity of 358.27 dB/RIU ranging from 1.33 to 1.38. The sensor has also been implemented for the concentration measurement of λ-phage DNA solution. In addition, the dip intensity is insensitive to the ambient temperature variation, making it a good candidate for temperature-independent bio-sensing area.

  2. Assessment of the degree of contamination of rat germ cell preparations using specific cDNA probes

    Directory of Open Access Journals (Sweden)

    Savaris R.F.

    1997-01-01

    Full Text Available Recent reports showing a decrease in sperm count in men have brought new concerns about male infertility. Animal models have been widely used to provide some relevant information about the human male gamete, and extrapolations are made to men and to the clinical context. The present study assesses one of the methods used for separation of germ cells of the adult rat testis, namely centrifugal elutriation followed by density gradients (Percoll®. This method was chosen since it presents the best results for cell purity in separating germ cells from the rat testis. A comparison between continuous and discontinuous Percoll® gradients was performed in order to identify the best type of gradient to separate the cells. Maximal cell purity was obtained for spermatocytes (81 ± 8.2%, mean ± SEM and spermatids (84 ± 2.6% using centrifugal elutriation followed by continuous Percoll® gradients. A significant difference in purity was observed between elongating spermatids harvested from continuous Percoll® gradients and from discontinuous gradients. Molecular analysis was used to assess cell contamination by employing specific probes, namely transition protein 2 (TP2, mitochondrial cytochrome C oxidase II (COX II, and sulfated glycoprotein 1 (SGP1. Molecular analysis of the samples demonstrated that morphological criteria are efficient in characterizing the main composition of the cell suspension, but are not reliable for identifying minimal contamination from other cells. Reliable cell purity data should be established using molecular analysis

  3. Micro-RNA detection based on fluorescence resonance energy transfer of DNA-carbon quantum dots probes.

    Science.gov (United States)

    Khakbaz, Faeze; Mahani, Mohamad

    2017-04-15

    Carbon quantum dots have been proposed as an effective platform for miRNA detection. Carbon dots were synthesized by citric acid. The synthesized dots were characterized by dynamic light scattering, UV-Vis spectrophotometry, spectrofluorimetry, transmission electron microscopy and FT-IR spectrophotometry. The fluorescence quantum yield of the synthesized dots was determined using quinine sulfate as the standard. The FAM-labeled single stranded DNA, as sensing element, was adsorbed on dots by π-π interaction. The quenching of the dots fluorescence due to fluorescence resonance energy transfer (FRET) was used for mir 9-1 detection. In the presence of the complementary miRNA, the FRET did not take place and the fluorescence was recovered.

  4. Use of cell-SELEX to generate DNA aptamers as molecular probes of HPV-associated cervical cancer cells.

    Science.gov (United States)

    Graham, Jessica C; Zarbl, Helmut

    2012-01-01

    Disease-specific biomarkers are an important tool for the timely and effective management of pathological conditions, including determination of susceptibility, diagnosis, and monitoring efficacy of preventive or therapeutic strategies. Aptamers, comprising single-stranded or double-stranded DNA or RNA, can serve as biomarkers of disease or biological states. Aptamers can bind to specific epitopes on macromolecules by virtue of their three dimensional structures and, much like antibodies, aptamers can be used to target specific epitopes on the basis of their molecular shape. The Systematic Evolution of Ligands by EXponential enrichment (SELEX) is the approach used to select high affinity aptamers for specific macromolecular targets from among the >10(13) oligomers comprising typical random oligomer libraries. In the present study, we used live cell-based SELEX to identify DNA aptamers which recognize cell surface differences between HPV-transformed cervical carcinoma cancer cells and isogenic, nontumorigenic, revertant cell lines. Whole-cell SELEX methodology was adapted for use with adherent cell lines (which we termed Adherent Cell-SELEX (AC-SELEX)). Using this approach, we identified high affinity aptamers (nanomolar range K(d)) to epitopes specific to the cell surface of two nontumorigenic, nontumorigenic revertants derived from the human cervical cancer HeLa cell line, and demonstrated the loss of these epitopes in another human papillomavirus transformed cervical cancer cell line (SiHa). We also performed preliminary investigation of the aptamer epitopes and their binding characteristics. Using AC-SELEX we have generated several aptamers that have high affinity and specificity to the nontumorigenic, revertant of HPV-transformed cervical cancer cells. These aptamers can be used to identify new biomarkers that are related to carcinogenesis. Panels of aptamers, such as these may be useful in predicting the tumorigenic potential and properties of cancer biopsies

  5. Use of cell-SELEX to generate DNA aptamers as molecular probes of HPV-associated cervical cancer cells.

    Directory of Open Access Journals (Sweden)

    Jessica C Graham

    Full Text Available BACKGROUND: Disease-specific biomarkers are an important tool for the timely and effective management of pathological conditions, including determination of susceptibility, diagnosis, and monitoring efficacy of preventive or therapeutic strategies. Aptamers, comprising single-stranded or double-stranded DNA or RNA, can serve as biomarkers of disease or biological states. Aptamers can bind to specific epitopes on macromolecules by virtue of their three dimensional structures and, much like antibodies, aptamers can be used to target specific epitopes on the basis of their molecular shape. The Systematic Evolution of Ligands by EXponential enrichment (SELEX is the approach used to select high affinity aptamers for specific macromolecular targets from among the >10(13 oligomers comprising typical random oligomer libraries. In the present study, we used live cell-based SELEX to identify DNA aptamers which recognize cell surface differences between HPV-transformed cervical carcinoma cancer cells and isogenic, nontumorigenic, revertant cell lines. METHODOLOGY/PRINCIPAL FINDINGS: Whole-cell SELEX methodology was adapted for use with adherent cell lines (which we termed Adherent Cell-SELEX (AC-SELEX. Using this approach, we identified high affinity aptamers (nanomolar range K(d to epitopes specific to the cell surface of two nontumorigenic, nontumorigenic revertants derived from the human cervical cancer HeLa cell line, and demonstrated the loss of these epitopes in another human papillomavirus transformed cervical cancer cell line (SiHa. We also performed preliminary investigation of the aptamer epitopes and their binding characteristics. CONCLUSIONS/SIGNIFICANCE: Using AC-SELEX we have generated several aptamers that have high affinity and specificity to the nontumorigenic, revertant of HPV-transformed cervical cancer cells. These aptamers can be used to identify new biomarkers that are related to carcinogenesis. Panels of aptamers, such as these may

  6. Mobile Probing and Probes

    DEFF Research Database (Denmark)

    2013-01-01

    Mobile probing is a method, developed for learning about digital work situations, as an approach to discover new grounds. The method can be used when there is a need to know more about users and their work with certain tasks, but where users at the same time are distributed (in time and space......). Mobile probing was inspired by the cultural probe method, and was influenced by qualitative interview and inquiry approaches. The method has been used in two subsequent projects, involving school children (young adults at 15-17 years old) and employees (adults) in a consultancy company. Findings point...... to mobile probing being a flexible method for uncovering the unknowns, as a way of getting rich data to the analysis and design phases. On the other hand it is difficult to engage users to give in depth explanations, which seem easier in synchronous dialogs (whether online or face2face). The development...

  7. Mobile Probing and Probes

    DEFF Research Database (Denmark)

    2012-01-01

    Mobile probing is a method, which has been developed for learning about digital work situations, as an approach to discover new grounds. The method can be used when there is a need to know more about users and their work with certain tasks, but where users at the same time are distributed (in time...... and space). Mobile probing was inspired by the cultural probe method, and was influenced by qualitative interview and inquiry approaches. The method has been used in two subsequent projects, involving school children (young adults at 15-17 years old) and employees (adults) in a consultancy company. Findings...... point to mobile probing being a flexible method for uncovering the unknowns, as a way of getting rich data to the analysis and design phases. On the other hand it is difficult to engage users to give in depth explanations, which seem easier in synchronous dialogs (whether online or face2face...

  8. Mobile Probing and Probes

    DEFF Research Database (Denmark)

    2012-01-01

    Mobile probing is a method, which has been developed for learning about digital work situations, as an approach to discover new grounds. The method can be used when there is a need to know more about users and their work with certain tasks, but where users at the same time are distributed (in time...... and space). Mobile probing was inspired by the cultural probe method, and was influenced by qualitative interview and inquiry approaches. The method has been used in two subsequent projects, involving school children (young adults at 15-17 years old) and employees (adults) in a consultancy company. Findings...... point to mobile probing being a flexible method for uncovering the unknowns, as a way of getting rich data to the analysis and design phases. On the other hand it is difficult to engage users to give in depth explanations, which seem easier in synchronous dialogs (whether online or face2face...

  9. Development of DNA probe for detection of Aujeszky's disease virus Desenvolvimento de uma sonda de DNA para a detecção do vírus da doença de Aujeszky

    Directory of Open Access Journals (Sweden)

    A.L. Cândido

    1999-08-01

    Full Text Available A DNA hybridization dot-blot assay using a radioactive and a non-radioactive probe has been developed for the detection of Aujeszky's disease virus (ADV. The Bam H I 7 fragment of ADV genomic DNA was labeled by nick translation using 32P-dCTP and the 196bp polymerase chain reaction (PCR gG glycoprotein gene amplified fragment was also labeled by nick translation but using biotin d-7-dATP. This technique provides a fast and effective means of detecting acute cases of ADV infection but it was unable to detect ADV nucleic acid sequences in trigeminal nerve ganglia of latent infected pigs and mice.Uma sonda radioativa e uma outra biotinilada foram produzidas para detecção do vírus da doença de Aujeszky (VDA. O fragmento de Bam H I 7 do DNA genômico do VDA foi marcado por "nick translation" empregando P32dCTP e um fragmento de 196pb, resultante de uma amplificação pela reação em cadeia pela polimerase marcado com biotina 7-dATP. Essas sondas, de uma maneira rápida e específica, prestaram-se para a detecção da infecção aguda pelo VDA. Entretanto, não se mostraram sensíveis o suficiente para detectar seqüências genômicas de VDA no gânglio trigêmio de suínos e camundongos com infecção latente.

  10. Diagnosis and quantification of glycerol assimilating denitrifying bacteria in an integrated fixed-film activated sludge reactor via 13C DNA stable-isotope probing.

    Science.gov (United States)

    Lu, Huijie; Chandran, Kartik

    2010-12-01

    Glycerol, a byproduct of biodiesel and oleo-chemical manufacturing operations, represents an attractive alternate to methanol as a carbon and electron donor for enhanced denitrification. However, unlike methanol, little is known about the diversity and activity of glycerol assimilating bacteria in activated sludge. In this study, the microbial ecology of glycerol assimilating denitrifying bacteria in a sequencing batch integrated fixed film activated sludge (SB-IFAS) reactor was investigated using (13)C-DNA stable isotope probing (SIP). During steady state SB-IFAS reactor operation, near complete nitrate removal (92.7 ± 5.8%) was achieved. Based on (13)C DNA clone libraries obtained after 360 days of SB-IFAS reactor operation, bacteria related to Comamonas spp. and Diaphorobacter spp. dominated in the suspended phase communities. (13)C assimilating members in the biofilm community were phylogenetically more diverse and were related to Comamonas spp., Bradyrhizobium spp., and Tessaracoccus spp. Possibly owing to greater substrate availability in the suspended phase, the glycerol-assimilating denitrifying populations (quantified by real-time PCR) were more abundant therein than in the biofilm phase. The biomass in the suspended phase also had a higher specific denitrification rate than the biofilm phase (p = 4.33e-4), and contributed to 69.7 ± 4.5% of the overall N-removal on a mass basis. The kinetics of glycerol based denitrification by suspended phase biomass were approximately 3 times higher than with methanol. Previously identified methanol assimilating denitrifying bacteria were not associated with glycerol assimilation, thereby suggesting limited cross-utilization of these two substrates for denitrification in the system tested.

  11. DNA stable-isotope probing of oil sands tailings pond enrichment cultures reveals different key players for toluene degradation under methanogenic and sulfidogenic conditions.

    Science.gov (United States)

    Laban, Nidal Abu; Dao, Anh; Foght, Julia

    2015-05-01

    Oil sands tailings ponds are anaerobic repositories of fluid wastes produced by extraction of bitumen from oil sands ores. Diverse indigenous microbiota biodegrade hydrocarbons (including toluene) in situ, producing methane, carbon dioxide and/or hydrogen sulfide, depending on electron acceptor availability. Stable-isotope probing of cultures enriched from tailings associated specific taxa and functional genes to (13)C6- and (12)C7-toluene degradation under methanogenic and sulfate-reducing conditions. Total DNA was subjected to isopycnic ultracentrifugation followed by gradient fraction analysis using terminal restriction fragment length polymorphism (T-RFLP) and construction of 16S rRNA, benzylsuccinate synthase (bssA) and dissimilatory sulfite reductase (dsrB) gene clone libraries. T-RFLP analysis plus sequencing and in silico digestion of cloned taxonomic and functional genes revealed that Clostridiales, particularly Desulfosporosinus (136 bp T-RF) contained bssA genes and were key toluene degraders during methanogenesis dominated by Methanosaeta. Deltaproteobacterial Desulfobulbaceae (157 bp T-RF) became dominant under sulfidogenic conditions, likely because the Desulfosporosinus T-RF 136 apparently lacks dsrB and therefore, unlike its close relatives, is presumed incapable of dissimilatory sulfate reduction. We infer incomplete oxidation of toluene by Desulfosporosinus in syntrophic association with Methanosaeta under methanogenic conditions, and complete toluene oxidation by Desulfobulbaceae during sulfate reduction. © FEMS 2015. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

  12. Polydopamine-Assisted Fabrication of Fiber-Optic Localized Surface Plasmon Resonance Sensor Based on Gold Nanoparticles

    Institute of Scientific and Technical Information of China (English)

    苏荣欣; 裴哲远; 黄仁亮; 齐崴; 王梦凡; 王利兵; 何志敏

    2015-01-01

    A fast and facile method of fabricating fiber-optic localized surface plasmon resonance sensors based on spherical gold nanoparticles was introduced in this study. The gold nanoparticles with an average diameter of 55 nm were synthesized via the Turkevich method and were then immobilized onto the surface of an uncladded sensor probe using a polydopamine layer. To obtain a sensor probe with high sensitivity to changes in the refractive index, a set of key optimization parameters, including the sensing length, coating time of the polydopamine layer, and coating time of the gold nanoparticles, were investigated. The sensitivity of the optimized sensor probe was 522.80 nm per refractive index unit, and the probe showed distinctive wavelength shifts when the refractive index was changed from 1.328 6 to 1.398,7. When stored in deionized water at 4℃, the sensor probe proved to be stable over a period of two weeks. The sensor also exhibited advantages, such as low cost, fast fabrication, and simple optical setup, which indicated its potential application in remote sensing and real-time detection.

  13. Neural Network-Based Multimode Fiber-Optic Information Transmission

    Science.gov (United States)

    Marusarz, Ronald K.; Sayeh, Mohammad R.

    2001-01-01

    A new technique for transmitting information through multimode fiber-optic cables is presented. This technique sends parallel channels through the fiber-optic cable, thereby greatly improving the data transmission rate compared with that of the current technology, which uses serial data transmission through single-mode fiber. An artificial neural network is employed to decipher the transmitted information from the received speckle pattern. Several different preprocessing algorithms are developed, tested, and evaluated. These algorithms employ average region intensity, distributed individual pixel intensity, and maximum mean-square-difference optimal group selection methods. The effect of modal dispersion on the data rate is analyzed. An increased data transmission rate by a factor of 37 over that of single-mode fibers is realized. When implementing our technique, we can increase the channel capacity of a typical multimode fiber by a factor of 6.

  14. [Value of flexible fiberoptic bronchoscopy under local anesthesia in infants].

    Science.gov (United States)

    Bodart, E; De Lange, M; Vliers, A

    1993-06-01

    From October 1991 through April 1992, 16 infants aged 5 to 25 months (mean age 14.3 months) underwent bronchoscopy with a flexible fiberoptic bronchoscope, under local anesthesia. The technique is described in detail. Reasons for bronchoscopy included recurrent or persistent pneumonia (n = 4), persistent atelectasia (n = 4), lymphadenopathy and/or airway compression (n = 2), suspected foreign body (n = 2), bronchoalveolar lavage to investigate diffuse interstitial lung disease (n = 2), and severe recurrent wheezing (n = 2). The procedure established the accurate diagnosis in 14 cases. Adverse events (32%) were minor (transient hypoxia, n = 3; moderate fever, n = 1; and laryngospasm, n = 1) and resolved completely. Flexible fiberoptic bronchoscopy under local anesthesia is a simple procedure which is safe in patients under 30 months of age when performed by a experienced operator in an adequate facility. This method is useful for the diagnosis and/or treatment of a broad spectrum of conditions.

  15. Mobile Probing and Probes

    DEFF Research Database (Denmark)

    2013-01-01

    to mobile probing being a flexible method for uncovering the unknowns, as a way of getting rich data to the analysis and design phases. On the other hand it is difficult to engage users to give in depth explanations, which seem easier in synchronous dialogs (whether online or face2face). The development...

  16. Review Of Fiber-Optic Electric-Field Sensors

    Science.gov (United States)

    De Paula, Ramon P.; Jarzynski, Jacek

    1989-01-01

    Tutorial paper reviews state of art in fiber-optic sensors of alternating electric fields. Because such sensors are made entirely of dielectric materials, they are relatively transparent to incident electric fields; they do not distort fields significantly. Paper presents equations that express relationships among stress, strain, and electric field in piezoactive plastic and equations for phase shift in terms of photoelastic coefficients and strains in optical fiber.

  17. Fiber-optic sensor applications in civil and geotechnical engineering

    Science.gov (United States)

    Habel, Wolfgang R.; Krebber, Katerina

    2011-09-01

    Different types of fiber-optic sensors based on glass or polymeric fibers are used to evaluate material behavior or to monitor the integrity and long-term stability of load-bearing structure components. Fiber-optic sensors have been established as a new and innovative measurement technology in very different fields, such as material science, civil engineering, light-weight structures, geotechnical areas as well as chemical and high-voltage substations. Very often, mechanical quantities such as deformation, strain or vibration are requested. However, measurement of chemical quantities in materials and structure components, such as pH value in steel reinforced concrete members also provides information about the integrity of concrete structures. A special fiber-optic chemical sensor for monitoring the alkaline state (pH value) of the cementitious matrix in steel-reinforced concrete structures with the purpose of early detection of corrosion-initiating factors is described. The paper presents the use of several fiber-optic sensor technologies in engineering. One example concerns the use of highly resolving concrete-embeddable fiber Fabry-Perot acoustic emission (AE) sensors for the assessment of the bearing behaviour of large concrete piles in existing foundations or during and after its installation. Another example concerns fiber Bragg grating (FBG) sensors attached to anchor steels (micro piles) to measure the strain distribution in loaded soil anchors. Polymer optical fibers (POF) can be — because of their high elasticity and high ultimate strain — well integrated into textiles to monitor their deformation behaviour. Such "intelligent" textiles are capable of monitoring displacement of soil or slopes, critical mechanical deformation in geotechnical structures (dikes, dams, and embankments) as well as in masonry structures during and after earthquakes.

  18. Fiber-Optic Micrometeoroid/Orbital Debris Impact Detector System

    Science.gov (United States)

    Christiansen, Eric L.; Tennyson, R. C.; Morison, W. D.

    2012-01-01

    A document describes a reliable, lightweight micrometeoroid/orbital debris (MMOD) detection system that can be located at strategic positions of "high consequence" to provide real-time warning of a penetration, its location, and the extent of the damage to a spacecraft. The concept is to employ fiber-optic sensors to detect impact damage and penetration of spacecraft structures. The fibers are non-electrical, employ light waves, and are immune to electromagnetic interference. The fiber-optic sensor array can be made as a stand-alone product, being bonded to a flexible membrane material or a structure that is employed as a MMOD shield material. The optical sensors can also be woven into hybrid MMOD shielding fabrics. The glass fibers of the fiber-optic sensor provide a dual purpose in contributing to the breakup of MMOD projectiles. The grid arrays can be made in a modular configuration to provide coverage over any area desired. Each module can be connected to a central scanner instrument and be interrogated in a continuous or periodic mode.

  19. Molecular DNA switches and DNA chips

    Science.gov (United States)

    Sabanayagam, Chandran R.; Berkey, Cristin; Lavi, Uri; Cantor, Charles R.; Smith, Cassandra L.

    1999-06-01

    We present an assay to detect single-nucleotide polymorphisms on a chip using molecular DNA switches and isothermal rolling- circle amplification. The basic principle behind the switch is an allele-specific oligonucleotide circularization, mediated by DNA ligase. A DNA switch is closed when perfect hybridization between the probe oligonucleotide and target DNA allows ligase to covalently circularize the probe. Mismatches around the ligation site prevent probe circularization, resulting in an open switch. DNA polymerase is then used to preferentially amplify the closed switches, via rolling-circle amplification. The stringency of the molecular switches yields 102 - 103 fold discrimination between matched and mismatched sequences.

  20. Miniature fiber-optic force sensor for vitreoretinal microsurgery based on low-coherence Fabry-Pérot interferometry

    Science.gov (United States)

    Liu, Xuan; Iordachita, Iulian I.; He, Xingchi; Taylor, Russell H.; Kang, Jin U.

    2012-01-01

    Vitreoretinal surgery requires delicate manipulation of retinal tissue. However, tool-to-tissue interaction forces in the order of sub-millinewton are usually below the human sensory threshold. A surgical force sensor (FS) compatible with conventional surgical tools may significantly improve the surgery outcome by preventing tissue damage. We have designed and built a miniature FS for vitreoretinal surgery using a fiber-optic common-path phase-sensitive optical coherence tomography (OCT) system where the distal end of the fiber probe forms a low-finesse Fabry-Pérot (FP) cavity between the cleaved tip of the lead-in single mode fiber and the polished back surface of a stainless steel surgical tool tip. To accurately measure the change of the FP cavity length, the cavity is interrogated by the fiber-optic common-path phase-sensitive OCT. The FP cavity was illuminated with a broadband light source, and the interferometric signal was detected using a broadband spectrometer. The phase of the interferometric signal, which is proportional to the cavity length change as well as the exerted force, was extracted. We have conducted calibration experiments to characterize our one dimensional FS. Our result shows that the FS responses linearly to force in axial direction with force sensitivity better than 0.25 millinewton.

  1. Synthesis of PET probe O(6)-[(3-[(11)C]methyl)benzyl]guanine by Pd(0)-mediated rapid C-[(11)C]methylation toward imaging DNA repair protein O(6)-methylguanine-DNA methyltransferase in glioblastoma.

    Science.gov (United States)

    Koyama, Hiroko; Ikenuma, Hiroshi; Toda, Hiroshi; Kondo, Goro; Hirano, Masaki; Kato, Masaya; Abe, Junichiro; Yamada, Takashi; Wakabayashi, Toshihiko; Ito, Kengo; Natsume, Atsushi; Suzuki, Masaaki

    2017-03-18

    O(6)-Benzylguanine (O(6)-BG) is a substrate of O(6)-methylguanine-DNA methyltransferase (MGMT), which is involved in drug resistance of chemotherapy in the majority of glioblastoma multiform. For clinical diagnosis, it is hoped that the MGMT expression level could be determined by a noninvasive method to understand the detailed biological properties of MGMT-specific tumors. We synthesized (11)C-labeled O(6)-[(3-methyl)benzyl]guanine ([(11)C]mMeBG) as a positron emission tomography probe. Thus, a mixed amine-protected stannyl precursor, N(9)-(tert-butoxycarbonyl)-O(6)-[3-(tributylstannyl)benzyl]-N(2)-(trifluoroacetyl)guanine, was subjected to rapid C-[(11)C]methylation under [(11)C]CH3I/[Pd2(dba)3]/P(o-CH3C6H4)3/CuCl/K2CO3 in NMP, followed by quick deprotection with LiOH/H2O, giving [(11)C]mMeBG with total radioactivity of 1.34GBq and ≥99% radiochemical and chemical purities.

  2. Human papillomavirus detection in cervical dysplasias or neoplasias and in condylomata acuminaata by in situ hybridization with biotinylated DNA probes Detecção de papilomavirus humano em displasias ou neoplasias cervicais e em condilomas acuminados por hibridização in situ com sondas de DNA biotiniladas

    OpenAIRE

    Eliane Machado Guimarães; Geraldo Brasileiro Filho; Sérgio Danilo Junho Pena

    1992-01-01

    Specimens from cervical dysplasias or carcinomas and genital condylomata acuminata were retrospectively analysed by in situ hybridization (ISH) with bioti-nylated DNA probes for human papillomavirus (HPV) types 6, 11, 16 and 18. In the control group no case was positive for HPV DNA. In mild/moderate dysplasias, 4 cases (14%) were positive for HPV 6 or 11 and 2 cases (7%), for HPV 16. In the severe dysplasia/in situ carcinoma group, 9 cases (31%) showed presence of DNA of HPV types 16 or 18. S...

  3. Compensation of chromatic and polarization mode dispersion in fiber-optic communication lines in microwave signals transmittion.

    Science.gov (United States)

    Ermolaev, A. N.; Krishpents, G. P.; Davydov, V. V.; Vysoczkiy, M. G.

    2016-08-01

    Methods of dispersion compensation in fiber-optic communication lines. A new proposed method of electronic dispersion compensation in the transmission of microwave signals through fiber-optic lines. Represents is proposed the results of experimental studies of this method.

  4. Characterization of Growing Soil Bacterial Communities across a pH gradient Using H218O DNA-Stable Isotope Probing

    Science.gov (United States)

    Welty-Bernard, A. T.; Schwartz, E.

    2014-12-01

    Recent studies have established consistent relationships between pH and bacterial diversity and community structure in soils from site-specific to landscape scales. However, these studies rely on DNA or PLFA extraction techniques from bulk soils that encompass metabolically active and inactive, or dormant, communities, and loose DNA. Dormant cells may comprise up to 80% of total live cells. If dormant cells dominate a particular environment, it is possible that previous interpretations of the soil variables assumed to drive communities could be profoundly affected. We used H218O stable isotope probing and bar-coded illumina sequencing of 16S rRNA genes to monitor the response of actively growing communities to changes in soil pH in a soil microcosm over 14 days. This substrate-independent approach has several advantages over 13C or 15N-labelled molecules in that all growing bacteria should be able to make use of water, allowing characterization of whole communities. We hypothesized that Acidobacteria would increasingly dominate the growing community and that Actinobacteria and Bacteroidetes would decline, given previously established responses by these taxa to soil pH. Instead, we observed the reverse. Actinobacteria abundance increased three-fold from 26 to 76% of the overall community as soil pH fell from pH 5.6 to pH 4.6. Shifts in community structure and decreases in diversity with declining soil pH were essentially driven by two families, Streptomyceaca and Microbacteracea, which collectively increased from 2 to 40% of the entire community. In contrast, Acidobacteria as a whole declined although numbers of subdivision 1 remained stable across all soil pH levels. We suggest that the brief incubation period in this SIP study selected for growth of acid-tolerant Actinobacteria over Acidobacteria. Taxa within Actinomycetales have been readily cultured over short time frames, suggesting rapid growth patterns. Conversely, taxa within Acidobacteria have been

  5. An inexpensive and simple method for thermally stable immobilization of DNA on an unmodified glass surface: UV linking of poly(T)10-poly(C)10-tagged DNA probes

    DEFF Research Database (Denmark)

    Guðnason, Haukur; Dufva, Hans Martin; Bang, Dang Duong;

    2008-01-01

    be linked by UV light irradiation onto a plain, unmodified glass surface. Probes immobilized onto unmodified glass microscope slides performed similarly to probes bound to commercial amino-silane-coated slides and had comparable detection limits. The TC-tagged probes linked to unmodified glass did not show...

  6. A new fiber-optic non-contact compact laser-ultrasound scanner for fast non-destructive testing and evaluation of aircraft composites.

    Science.gov (United States)

    Pelivanov, Ivan; Buma, Takashi; Xia, Jinjun; Wei, Chen-Wei; O'Donnell, Matthew

    2014-03-21

    Laser ultrasonic (LU) inspection represents an attractive, non-contact method to evaluate composite materials. Current non-contact systems, however, have relatively low sensitivity compared to contact piezoelectric detection. They are also difficult to adjust, very expensive, and strongly influenced by environmental noise. Here, we demonstrate that most of these drawbacks can be eliminated by combining a new generation of compact, inexpensive fiber lasers with new developments in fiber telecommunication optics and an optimally designed balanced probe scheme. In particular, a new type of a balanced fiber-optic Sagnac interferometer is presented as part of an all-optical LU pump-probe system for non-destructive testing and evaluation of aircraft composites. The performance of the LU system is demonstrated on a composite sample with known defects. Wide-band ultrasound probe signals are generated directly at the sample surface with a pulsed fiber laser delivering nanosecond laser pulses at a repetition rate up to 76 kHz rate with a pulse energy of 0.6 mJ. A balanced fiber-optic Sagnac interferometer is employed to detect pressure signals at the same point on the composite surface. A- and B-scans obtained with the Sagnac interferometer are compared to those made with a contact wide-band polyvinylidene fluoride transducer.

  7. Grazing-angle fiber-optic fourier transform infrared reflection-absorption spectroscopy for the in situ detection and quantification of two active pharmaceutical ingredients on glass.

    Science.gov (United States)

    Perston, Benjamin B; Hamilton, Michelle L; Williamson, Bryce E; Harland, Peter W; Thomson, Mary A; Melling, Peter J

    2007-02-01

    Fourier transform infrared reflection-absorption spectroscopy has been used with a fiber-optic grazing-angle reflectance probe as a rapid, in situ method for trace surface analysis of acetaminophen and aspirin at loadings of approximately 0-2 microg cm(-2) on glass. Partial least-squares multivariate regression permits the loadings to be quantified, simultaneously, with root-mean-squared errors of prediction of RMSEP approximately 0.1 microg cm(-2) for both compounds. The detection limits are estimated to be LD approximately 0.2 microg cm(-2).

  8. 用于MLPA技术的单链长探针制备方法研究%A novel procedure for the preparation of the downstream long single-strand DNA probes for multiplex ligation-dependent probe amplification

    Institute of Scientific and Technical Information of China (English)

    陈菲; 陈枝楠; 康林; 潘广; 叶奕优; 肖启明; 凌杏园

    2012-01-01

    依据多重连接依赖探针扩增技术(MLPA)的单链探针设计要求,将检测各基因位点的MPLA探针对的上游探针设计为短探针,采用化学法加以合成;将下游探针作为长探针,以不对称PCR方法制备。本文应用该技术制备了检测12个转基因玉米品系的MLPA下游探针;并用其中的2个长探针进行转基因玉米品系检测,结果表明制备探针完全符合转基因检测要求。该技术操作简单,成本低,应用价值高。%According to the design principles of single-stranded probe in MLPA,each upstream probe of the MLPA probe pairs for different loci are designed as short probes and synthesized chemically,however,the downstream probes are designed as long oligo-nucleotides probes and prepared with asymmetric PCR.In this study,With this novel procedure,12 such probes for 12 GMO corn lines were prepared,and were proved to be effective with 2 such long probes being practically used for GMO detection.This novel procedure has the characteristic of simple manipulation and low cost,thus has high value for practical use.

  9. A signal-on electrochemical DNA biosensor based on potential-assisted Cu(I)-catalyzed azide-alkyne cycloaddition mediated labeling of hairpin-like oligonucleotide with electroactive probe.

    Science.gov (United States)

    Hu, Qiong; Kong, Jinming; Li, Yajie; Zhang, Xueji

    2016-01-15

    A novel electrochemical biosensor was developed for the signal-on detection of sequence-specific DNA by exploiting potential-assisted Cu(I)-catalyzed azide-alkyne cycloaddition (φCuAAC) as an efficient approach for the labeling of hairpin-like oligonucleotide (hairpin) with electroactive probe. The hairpins, dually labeled with thiol and azide at either terminal, were firstly self-assembled on gold electrode and served as the capture probes for the specific recognition of target DNA. Upon hybridization with target DNA, the surface-confined hairpins were unfolded, liberating the azide-containing terminals away from electrode surface. Subsequently, the unfolded hairpins were conveniently and efficiently labeled with ethynylferrocene (EFC) via the φCuAAC. The quantitatively labeled EFC was finally measured via differential pulse voltammetry (DPV) for the signal-on electrochemical detection of sequence-specific DNA. The biosensor presented a good linear response over the range from 1pM to 1nM with a detection limit of 0.62pM. Results also revealed that it was highly specific and held a good detection capability in serum samples. Furthermore, the ability to chemoselectively label hairpin-like oligonucleotide with signal reporter by electrical addressing, together with the simplicity and efficiency of the φCuAAC, makes it compatible with microfluidic devices and microelectrode arrays to achieve the miniaturized and multiplexed detections.

  10. 21 CFR 874.4350 - Ear, nose, and throat fiberoptic light source and carrier.

    Science.gov (United States)

    2010-04-01

    ... 21 Food and Drugs 8 2010-04-01 2010-04-01 false Ear, nose, and throat fiberoptic light source and... HUMAN SERVICES (CONTINUED) MEDICAL DEVICES EAR, NOSE, AND THROAT DEVICES Surgical Devices § 874.4350 Ear, nose, and throat fiberoptic light source and carrier. (a) Identification. An ear, nose, and...

  11. Feasibility study on fiber-optic goniometer for measuring knee joint angle

    Science.gov (United States)

    Kim, Seon Geun; Jang, Kyoung Won; Yoo, Wook Jae; Shin, Sang Hun; Cho, Seunghyun; Lee, Bongsoo

    2014-09-01

    In this study, we fabricated a fiber-optic goniometer using a plastic optical fiber, a light-emitting diode, and a photodiode. The cladding of the plastic optical fiber was removed at 0.5-3mm intervals regularly to increase the bending loss of the plastic optical fiber. Also, the output voltages of the photodiode based on light intensity that measured using the fiber-optic goniometer were measured light intensities using the fiber-optic goniometer were measured to evaluate the linearity of the fiber-optic goniometer. Finally, we measured the responses of the fiber-optic goniometer for gait speeds of 3, 5, and 10 km/h using a data acquisition board and a LabVIEW program.

  12. CATV Fiberoptic Cable Markets In The U.S.

    Science.gov (United States)

    Kessler, John N.

    1984-03-01

    "Bandwidth on a metropolitan CATV cable plant is a resource that may be compared today with the crude oil beneath the Arabian desert at the turn of the century. It will provide a radically new two-way communications medium for homes and businesses, and it will change not only the way we communicate but possibly even the way we live." This prediction was made last year by William Zachman, vice president of research for International Data Corp. If Zachman is right, and our research tends to indicate he is, then fiberoptics will become the transmission medium that will facilitate two-way broadband communication. However, many of the multiple system CATV operators in the U.S. as well as many of the manufacturers of fiberoptic waveguide and cable believe that interactive video systems are not really very important. They are wrong. Broadband fiberoptic systems have just begun to impact the U.S. communications market. And within 5 years, tremendous changes will occur not only with regard to growth, but with regard to the supplier industry structure. Interactive, switched, video systems in the U.S. go back to the early 1960s, and the experiments at Bell Laboratories with what was then called Picturephone. This was com-pressed video transmission via telephone lines from one subscriber to another. Field trials were conducted in several U.S. cities for a period of years. But the commercial response to Picturephone at that time was so lacking that AT&T discontinued the trials and the work. But that is changing. Part of the reason for the change is due to the changing communication needs of the U.S. and of the world: the increase in the amount of information used by large numbers of people, the concurrent rise in the use of computers, the digitization of communications media, the rise in the level of experience with electronic communications, and perhaps most importantly the convergence of audio, data, image and video communications. There are technological reasons for the slow

  13. Fiber-Optic Monitoring System of Particle Counters

    Directory of Open Access Journals (Sweden)

    A. A. Titov

    2016-01-01

    Full Text Available The article considers development of a fiber-optic system to monitor the counters of particles. Presently, optical counters of particles, which are often arranged at considerable distance from each other, are used to study the saltation phenomenon. For monitoring the counters, can be used electric communication lines.However, it complicates and raises the price of system Therefore, we offered a fiber-optic system and the counter of particles, free from these shortcomings. The difference between the offered counter of particles and the known one is that the input of radiation to the counter and the output of radiation scattering on particles are made by the optical fibers, and direct radiation is entered the optical fiber rather than is delayed by a light trap and can be used for lighting the other counters thereby allowing to use their connection in series.The work involved a choice of the quartz multimode optical fiber for communication, defining the optical fiber and lenses parameters of the counter of particles, and a selection of the radiation source and the photo-detector.Using the theory of light diffraction on a particle, a measuring range of the particle sizes has been determined. The system speed has been estimated, and it has been shown that a range of communication can reach 200km.It should be noted that modulation noise of counters of particles connected in series have the impact on the useful signal. To assess the extent of this influence we have developed a calculation procedure to illustrate that with ten counters connected in series this influence on the signal-to-noise ratio will be insignificant.Thus, it has been shown that the offered fiber-optic system can be used for monitoring the counters of particles across the desertified territories. 

  14. Fiber-Optic Network Architectures for Onboard Avionics Applications Investigated

    Science.gov (United States)

    Nguyen, Hung D.; Ngo, Duc H.

    2003-01-01

    This project is part of a study within the Advanced Air Transportation Technologies program undertaken at the NASA Glenn Research Center. The main focus of the program is the improvement of air transportation, with particular emphasis on air transportation safety. Current and future advances in digital data communications between an aircraft and the outside world will require high-bandwidth onboard communication networks. Radiofrequency (RF) systems, with their interconnection network based on coaxial cables and waveguides, increase the complexity of communication systems onboard modern civil and military aircraft with respect to weight, power consumption, and safety. In addition, safety and reliability concerns from electromagnetic interference between the RF components embedded in these communication systems exist. A simple, reliable, and lightweight network that is free from the effects of electromagnetic interference and capable of supporting the broadband communications needs of future onboard digital avionics systems cannot be easily implemented using existing coaxial cable-based systems. Fiber-optical communication systems can meet all these challenges of modern avionics applications in an efficient, cost-effective manner. The objective of this project is to present a number of optical network architectures for onboard RF signal distribution. Because of the emergence of a number of digital avionics devices requiring high-bandwidth connectivity, fiber-optic RF networks onboard modern aircraft will play a vital role in ensuring a low-noise, highly reliable RF communication system. Two approaches are being used for network architectures for aircraft onboard fiber-optic distribution systems: a hybrid RF-optical network and an all-optical wavelength division multiplexing (WDM) network.

  15. Development of plasma bolometers using fiber-optic temperature sensors

    Science.gov (United States)

    Reinke, M. L.; Han, M.; Liu, G.; van Eden, G. G.; Evenblij, R.; Haverdings, M.; Stratton, B. C.

    2016-11-01

    Measurements of radiated power in magnetically confined plasmas are important for exhaust studies in present experiments and expected to be a critical diagnostic for future fusion reactors. Resistive bolometer sensors have long been utilized in tokamaks and helical devices but suffer from electromagnetic interference (EMI). Results are shown from initial testing of a new bolometer concept based on fiber-optic temperature sensor technology. A small, 80 μm diameter, 200 μm long silicon pillar attached to the end of a single mode fiber-optic cable acts as a Fabry-Pérot cavity when broadband light, λo ˜ 1550 nm, is transmitted along the fiber. Changes in temperature alter the optical path length of the cavity primarily through the thermo-optic effect, resulting in a shift of fringes reflected from the pillar detected using an I-MON 512 OEM spectrometer. While initially designed for use in liquids, this sensor has ideal properties for use as a plasma bolometer: a time constant, in air, of ˜150 ms, strong absorption in the spectral range of plasma emission, immunity to local EMI, and the ability to measure changes in temperature remotely. Its compact design offers unique opportunities for integration into the vacuum environment in places unsuitable for a resistive bolometer. Using a variable focus 5 mW, 405 nm, modulating laser, the signal to noise ratio versus power density of various bolometer technologies are directly compared, estimating the noise equivalent power density (NEPD). Present tests show the fiber-optic bolometer to have NEPD of 5-10 W/m2 when compared to those of the resistive bolometer which can achieve coatings, along with improving the spectral resolution of the interrogator.

  16. Modeling of Nonlinear Signal Distortion in Fiber-Optical Networks

    CERN Document Server

    Johannisson, Pontus

    2013-01-01

    A low-complexity model for signal quality prediction in a nonlinear fiber-optical network is developed. The model, which builds on the Gaussian noise model, takes into account the signal degradation caused by a combination of chromatic dispersion, nonlinear signal distortion, and amplifier noise. The center frequencies, bandwidths, and transmit powers can be chosen independently for each channel, which makes the model suitable for analysis and optimization of resource allocation, routing, and scheduling in large-scale optical networks applying flexible-grid wavelength-division multiplexing.

  17. Sampling impairments influence over stealthy fiber-optic signal decryption

    Science.gov (United States)

    Yeminy, Tomer; Sadot, Dan; Zalevsky, Zeev

    2013-03-01

    Recently we have proposed a method for temporal and spectral stealthy fiber-optic communications. In this technique the signal is spread in the time domain below the noise level and the power spectral density of the signal is spread beneath the noise level in the frequency domain. The power spectral density of the signal is reconstructed by means of sampling which results in coherent addition of the encrypted signal spectral replicas. Hence, the sampling quality plays a major role in the ability to successfully decrypt the received covert signal. In this paper, we mathematically and numerically analyze the effect of sampling impairments over the decryption performance.

  18. Fiber-Optic Gratings for Lidar Measurements of Water Vapor

    Science.gov (United States)

    Vann, Leila B.; DeYoung, Russell J.

    2006-01-01

    Narrow-band filters in the form of phase-shifted Fabry-Perot Bragg gratings incorporated into optical fibers are being developed for differential-absorption lidar (DIAL) instruments used to measure concentrations of atmospheric water vapor. The basic idea is to measure the relative amounts of pulsed laser light scattered from the atmosphere at two nearly equal wavelengths, one of which coincides with an absorption spectral peak of water molecules and the other corresponding to no water vapor absorption. As part of the DIAL measurement process, the scattered light is made to pass through a filter on the way to a photodetector. Omitting other details of DIAL for the sake of brevity, what is required of the filter is to provide a stop band that: Surrounds the water-vapor spectral absorption peaks at a wavelength of 946 nm, Has a spectral width of at least a couple of nanometers, Contains a pass band preferably no wider than necessary to accommodate the 946.0003-nm-wavelength water vapor absorption peak [which has 8.47 pm full width at half maximum (FWHM)], and Contains another pass band at the slightly shorter wavelength of 945.9 nm, where there is scattering of light from aerosol particles but no absorption by water molecules. Whereas filters used heretofore in DIAL have had bandwidths of =300 pm, recent progress in the art of fiber-optic Bragg-grating filters has made it feasible to reduce bandwidths to less than or equal to 20 pm and thereby to reduce background noise. Another benefit of substituting fiber-optic Bragg-grating filters for those now in use would be significant reductions in the weights of DIAL instruments. Yet another advantage of fiber-optic Bragg-grating filters is that their transmission spectra can be shifted to longer wavelengths by heating or stretching: hence, it is envisioned that future DIAL instruments would contain devices for fine adjustment of transmission wavelengths through stretching or heating of fiber-optic Bragg-grating filters

  19. Development of plasma bolometers using fiber-optic temperature sensors

    Energy Technology Data Exchange (ETDEWEB)

    Reinke, M. L., E-mail: reinkeml@ornl.gov [Oak Ridge National Laboratory, Oak Ridge, Tennessee 37831 (United States); Han, M.; Liu, G. [University of Nebraska-Lincoln, Lincoln, Nebraska 68588 (United States); Eden, G. G. van [Dutch Institute for Fundamental Energy Research, De Zaale 20, 5612 AJ Eindhoven (Netherlands); Evenblij, R.; Haverdings, M. [Technobis, Pyrietstraat 2, 1812 SC Alkmaar (Netherlands); Stratton, B. C. [Princeton Plasma Physics Laboratory, P.O. Box 451, Princeton, New Jersey 08543 (United States)

    2016-11-15

    Measurements of radiated power in magnetically confined plasmas are important for exhaust studies in present experiments and expected to be a critical diagnostic for future fusion reactors. Resistive bolometer sensors have long been utilized in tokamaks and helical devices but suffer from electromagnetic interference (EMI). Results are shown from initial testing of a new bolometer concept based on fiber-optic temperature sensor technology. A small, 80 μm diameter, 200 μm long silicon pillar attached to the end of a single mode fiber-optic cable acts as a Fabry–Pérot cavity when broadband light, λ{sub o} ∼ 1550 nm, is transmitted along the fiber. Changes in temperature alter the optical path length of the cavity primarily through the thermo-optic effect, resulting in a shift of fringes reflected from the pillar detected using an I-MON 512 OEM spectrometer. While initially designed for use in liquids, this sensor has ideal properties for use as a plasma bolometer: a time constant, in air, of ∼150 ms, strong absorption in the spectral range of plasma emission, immunity to local EMI, and the ability to measure changes in temperature remotely. Its compact design offers unique opportunities for integration into the vacuum environment in places unsuitable for a resistive bolometer. Using a variable focus 5 mW, 405 nm, modulating laser, the signal to noise ratio versus power density of various bolometer technologies are directly compared, estimating the noise equivalent power density (NEPD). Present tests show the fiber-optic bolometer to have NEPD of 5-10 W/m{sup 2} when compared to those of the resistive bolometer which can achieve <0.5 W/m{sup 2} in the laboratory, but this can degrade to 1-2 W/m{sup 2} or worse when installed on a tokamak. Concepts are discussed to improve the signal to noise ratio of this new fiber-optic bolometer by reducing the pillar height and adding thin metallic coatings, along with improving the spectral resolution of the interrogator.

  20. Field trial of composite fiber-optic overhead ground wire

    Science.gov (United States)

    Kubota, S.; Kawahira, H.; Nakajima, T.; Matsubara, I.; Saito, Y.; Kitayama, Y.

    A composite fiber-optic ground wire (OPGW), which provides additional communication capabilities for system protection and control of overhead power transmission systems has been developed. After laboratory tests, the OPGW was strung along a live power transmission line in a mountainous region and has been confirmed to have sufficient performance to establish a high-speed digital transmission network able to withstand actual conditions. The field line, constructed substantially by existing techniques, has proved that the new OPGW, accessories such as clamps and joint boxes, installation technique, and on-tower splicing method can be effectively utilized to produce a protection and control system with extremely stable characteristics.

  1. Fiber-optic fluorescence-quenching oxygen partial pressure sensor using platinum octaethylporphyrin.

    Science.gov (United States)

    Davenport, John J; Hickey, Michelle; Phillips, Justin P; Kyriacou, Panayiotis A

    2016-07-20

    The development and bench testing of a fiber-optic oxygen sensor is described. The sensor is designed for measurement of tissue oxygen levels in the mucosa of the digestive tract. The materials and construction are optimized for insertion through the mouth for measurement in the lower esophagus. An oxygen-sensitive fluorescence-quenching film was applied as a solution of platinum octaethylporphyrin (PtOEP) poly(ethyl methacrylate) (PEMA) and dichloromethane and dip coated onto the distal tip of the fiber. The sensor was tested by comparing relative fluorescence when immersed in liquid water at 37°C, at a range of partial pressures (0-101 kPa). Maximum relative fluorescence at most oxygen concentrations was seen when the PtOEP concentration was 0.1  g.L-1, four layers of coating solution were applied, and a fiber core radius of 600 μm was selected, giving a Stern-Volmer constant of 0.129  kPa-1. The performance of the sensor is suitable for many in vivo applications, particularly mucosal measurements. It has sufficient sensitivity, is sterilizable, and is sufficiently flexible and robust for insertion via the mouth without damage to the probe or risk of harm to the patient.

  2. Orientation-dependent fiber-optic accelerometer based on grating inscription over fiber cladding.

    Science.gov (United States)

    Rong, Qiangzhou; Qiao, Xueguang; Guo, Tuan; Bao, Weijia; Su, Dan; Yang, Hangzhou

    2014-12-01

    An orientation-sensitive fiber-optic accelerometer based on grating inscription over fiber cladding has been demonstrated. The sensor probe comprises a compact structure in which a short section of thin-core fiber (TCF) stub containing a "cladding" fiber Bragg grating (FBG) is spliced to another single-mode fiber (SMF) without any lateral offset. A femtosecond laser side-illumination technique was utilized to ensure that the grating inscription remains close to the core-cladding interface of the TCF. The core mode and the cladding mode of the TCF are coupled at the core-mismatch junction, and two well-defined resonances in reflection appear from the downstream FBG, in which the cladding resonance exhibits a strong polarization and bending dependence due to the asymmetrical distribution of the cladding FBG along the fiber cross section. Strong orientation dependence of the vibration (acceleration) measurement has been achieved by power detection of the cladding resonance. Meanwhile, the unwanted power fluctuations and temperature perturbations can be referenced out by monitoring the fundamental core resonance.

  3. Highly sensitive fiber-optic accelerometer by grating inscription in specific core dip fiber.

    Science.gov (United States)

    Rong, Qiangzhou; Guo, Tuan; Bao, Weijia; Shao, Zhihua; Peng, Gang-Ding; Qiao, Xueguang

    2017-09-19

    A highly sensitive fiber-optic accelerometer based on detecting the power output of resonances from the core dip is demonstrated. The sensing probe comprises a compact structure, hereby a short section of specific core (with a significant core dip) fiber stub containing a straight fiber Bragg grating is spliced to another single-mode fiber via a core self-alignment process. The femtosecond laser side-illumination technique was utilized to ensure that the grating inscription region is precisely positioned and compact in size. Two well-defined core resonances were achieved in reflection: one originates from the core dip and the other originates from fiber core. The key point is that only one of these two reflective resonances exhibits a high sensitivity to fiber bend (and vibration), whereas the other is immune to it. For low frequency (core mode reflection. Moreover, the sensor simultaneously provides an inherent power reference to eliminate unwanted power fluctuations from the light source and transmission lines, thus providing a means of evaluating weak seismic wave at low frequency.

  4. Spectroscopic investigation of the binding interactions of a membrane potential molecule in various supramolecular confined environments: contrasting behavior of surfactant molecules in relocation or release of the probe between nanocarriers and DNA surface.

    Science.gov (United States)

    Ghosh, Surajit; Banik, Debasis; Roy, Arpita; Kundu, Niloy; Kuchlyan, Jagannath; Sarkar, Nilmoni

    2014-12-01

    The fluorescence and optical properties of membrane potential probes are widely used to measure cellular transmembrane potentials. Hemicyanine dyes are also able to bind to membranes. The spectral properties of these molecules depend upon the charge shift from the donor moiety to the acceptor moiety. Changes in their spectral properties, i.e. absorption and emission maxima or intensities, are helpful in characterizing model membranes, microheterogeneous media, etc. In this article, we have demonstrated the binding interaction of a membrane potential probe, 1-ethyl-2-(4-(p-dimethylaminophenyl)-1,3-butadienyl)-pyridinium perchlorate (LDS 698), with various supramolecular confined environments. The larger dipole moment in the ground state compared to the excited state is a unique feature of hemicyanine dyes. Due to this unique feature, red shifts in the absorption maxima are observed in hydrophobic environments, compared with bulk solvent. On addition of surfactants and CT DNA to an aqueous solution containing LDS 698, significant increase in the emission intensity along with the quantum yield and lifetime indicate partition of the probe molecules into organized assemblies. In the case of the sodium dodecyl sulfate (SDS)-water system, due to interactions between the cationic LDS 698 and the anionic dodecyl sulfate moiety, the fluorescence intensity at ∼666 nm decreases and an additional peak at ∼590 nm appears at premicellar concentration (∼0.20 mM-4.50 mM). But at ∼5.50 mM SDS concentration, the absorbance in the higher wavelength region increases again, indicating encapsulation of the probe in micellar aggregates. This observation indicates that the premicellar aggregation behavior of SDS can also be judged by observing the changes in the UV-vis and fluorescence spectral patterns. The temperature dependent study also indicates that non-radiative deactivation of the dye molecules is highly restricted in the DNA micro-environment, compared with micelles

  5. Detection of mutations using microarrays of poly(C)10-poly(T)10 modified DNA probes immobilized on agarose films

    DEFF Research Database (Denmark)

    Dufva, Hans Martin; Petersen, Jesper; Stoltenborg, M.

    2006-01-01

    to an agarose film grafted onto unmodified glass. Microarrays of TC-tagged probes immobilized on the agarose film can be used to diagnose Mutations in the human P-globin gene, which encodes the beta-chains in hemoglobin. Although the probes differed widely regarding inciting point temperature (similar to 20...... degrees C), a single stringency wash still gave sufficiently high discrimination signals between perfect match and mismatch probes to allow robust mutation detection. In all, 270 genotypings were performed on patient materials, and no genotype was incorrectly classified. Quality control experiments...

  6. Probing the Salt Concentration Dependent Nucelobase Distribution in a Single-Stranded DNA-Single-Walled Carbon Nanotube Hybrid with Molecular Dynamics.

    Science.gov (United States)

    Ghosh, Soumadwip; Patel, Nisheet; Chakrabarti, Rajarshi

    2016-01-28

    The hybrids of single-walled carbon nanotube (SWCNT) and single stranded DNA (ssDNA) are novel nanoscale materials having remarkable applications in nanotechnology. The absorption of nucleobases on the surface of a SWCNT depends strongly on the ionic strength of the medium. In this paper, using atomistic molecular dynamics we have shown that at low salt concentration ssDNA wraps on the surface of SWCNT through hydrophobic π-π stacking between the DNA bases and the sp(2)-hybridized carbon atoms of the carbon nanotube. At high salt concentration, however, the DNA molecule adopts a partially folded structure and the ssDNA-SWCNT wrapping gets weakened significantly due to the self-stacking of the DNA bases. Our study can find relevance in CNT mediated gene delivery processes where subsequent unwrapping of the gene from its carrier is anticipated across the cell membrane regulated by an existing salt concentration gradient.

  7. Fiber-optic analog-to-NRZ binary conversion

    Science.gov (United States)

    Siahmakoun, A.; Reeves, E.

    2015-03-01

    A novel photonic analog-to-binary converter based on the first-order asynchronous delta-sigma modulation (ADSM) has been theoretically investigated and experimentally demonstrated. A fiber-optic prototype ADSM system is constructed and characterized. Delta-sigma modulation is a straightforward approach to A/D conversion because in this case an external clocking is not required and demodulation can be simply performed via a low-pass filtering process. To improve signal-to-noise ratio and thus system ENOB, a non-interferometric optical implementation has been constructed. The ADSM is comprised of three photonic devices: an inverted output photonic leaky integrator, bistable quantizer, and positive corrective feedback. The photonic integrator which is a recirculating loop performs the oversampling of an analog input using the cross-gain modulation in an SOA. We will show that the photonic ADSM produces an inverted non-return-to-zero (NRZ) pulse-density modulated output describing an input analog signal. This fiber-optic ADSM converts up to 7.6 MHz analog input at about 30 MS/s and effective ENOB of 6.

  8. Low-cost fiber-optic chemochromic hydrogen detector

    Energy Technology Data Exchange (ETDEWEB)

    Benson, D.K.; Tracy, C.E.; Hishmeh, G.; Ciszek, P.; Lee, S.H. [National Renewable Energy Lab., Golden, CO (United States)

    1998-08-01

    The ability to detect hydrogen gas leaks economically and with inherent safety is an important technology that could facilitate commercial acceptance of hydrogen fuel in various applications. In particular, hydrogen fueled passenger vehicles will require hydrogen leak detectors to signal the activation of safety devices such as shutoff valves, ventilating fans, alarms, etc. Such detectors may be required in several locations within a vehicle--wherever a leak could pose a safety hazard. It is therefore important that the detectors be very economical. This paper reports progress on the development of low-cost fiber-optic hydrogen detectors intended to meet the needs of a hydrogen-fueled passenger vehicle. In the design, the presence of hydrogen in air is sensed by a thin-film coating at the end of a polymer optical fiber. When the coating reacts reversibly with the hydrogen, its optical properties are changed. Light from a central electro-optic control unit is projected down the optical fiber where it is reflected from the sensor coating back to central optical detectors. A change in the reflected intensity indicates the presence of hydrogen. The fiber-optic detector offers inherent safety by removing all electrical power from the leak sites and offers reduced signal processing problems by minimizing electromagnetic interference. Critical detector performance requirements include high selectivity, response speed and durability as well as potential for low-cost production.

  9. Endotracheal Administration of Sufentanil and Tetracaine During Awake Fiberoptic Intubation.

    Science.gov (United States)

    Ji, Meng; Tao, Jun; Cheng, Min; Wang, Qingli

    2016-01-01

    Combined use of local anesthetics and low-dose opioids enhances the effects of local anesthetics. This study aimed to evaluate the efficacy of combined administration of sufentanil and tetracaine through the cricothyroid membrane during awake nasal intubation using fiberoptic bronchoscopy in patients with difficult airways. Forty patients were divided into 2 groups: group A received endotracheal administration of 25 μg of sufentanil and 2 mL of 1% tetracaine mixture; group B received endotracheal administration of 2 mL 1% tetracaine and routine local anesthetic sprays followed by slow intravenous injection of 25 μg of sufentanil. The results showed that endotracheal intubation was safely completed in all patients and vital signs including blood pressure, heart rate, and pulse oxygen saturation were not significantly different between groups A and B. However, time required for local anesthesia to take effect, time required to complete intubation, cough reflex, patient tolerance during intubation, and hemodynamic indices were significantly better in group A than in group B. In conclusion, our results suggest that endotracheal administration of sufentanil combined with tetracaine is safe, effective, and feasible in the context of awake nasal intubation using fiberoptic bronchoscopy.

  10. Computational imaging through a fiber-optic bundle

    Science.gov (United States)

    Lodhi, Muhammad A.; Dumas, John Paul; Pierce, Mark C.; Bajwa, Waheed U.

    2017-05-01

    Compressive sensing (CS) has proven to be a viable method for reconstructing high-resolution signals using low-resolution measurements. Integrating CS principles into an optical system allows for higher-resolution imaging using lower-resolution sensor arrays. In contrast to prior works on CS-based imaging, our focus in this paper is on imaging through fiber-optic bundles, in which manufacturing constraints limit individual fiber spacing to around 2 μm. This limitation essentially renders fiber-optic bundles as low-resolution sensors with relatively few resolvable points per unit area. These fiber bundles are often used in minimally invasive medical instruments for viewing tissue at macro and microscopic levels. While the compact nature and flexibility of fiber bundles allow for excellent tissue access in-vivo, imaging through fiber bundles does not provide the fine details of tissue features that is demanded in some medical situations. Our hypothesis is that adapting existing CS principles to fiber bundle-based optical systems will overcome the resolution limitation inherent in fiber-bundle imaging. In a previous paper we examined the practical challenges involved in implementing a highly parallel version of the single-pixel camera while focusing on synthetic objects. This paper extends the same architecture for fiber-bundle imaging under incoherent illumination and addresses some practical issues associated with imaging physical objects. Additionally, we model the optical non-idealities in the system to get lower modelling errors.

  11. Mono and trimethine cyanines Cyan 40 and Cyan 2 as probes for highly selective fluorescent detection of non-canonical DNA structures.

    Science.gov (United States)

    Kovalska, Vladyslava B; Losytskyy, Mykhaylo Yu; Yarmoluk, Sergiy M; Lubitz, Irit; Kotlyar, Alexander B

    2011-01-01

    Two of earlier reported dsDNA sensitive cyanine dyes-monomethine Cyan 40 and meso-substituted trimethine Cyan 2 were studied for their ability to interact with non-canonical DNA conformations. These dyes were characterized by spectral-luminescent methods in the presence of G-quadruplex, triplex and dsDNA motifs. We have demonstrated that Cyan 2 binds strongly and preferentially to triple- and quadruple-stranded DNA forms that results in a strong enhancement of the dye fluorescence, as compared to dsDNA, while Cyan 40 form fluorescent complexes preferentially only with the triplex form. Highly fluorescent complexes of Cyan 2 with DNA triplexes and G-quadruplexes and Cyan 40 with DNA triplexes are very stable and do not dissociate during gel electrophoresis, leading to preferential staining of the above DNA forms in gels. The data presented point to the intercalation mechanism of the Cyan 2 binding to G4-DNA, while the complexes of Cyan 40 and Cyan 2 with triplex DNA are believed to be formed via groove binding mode. The Cyan dyes can provide a highly sensitive method for detection and quantification of non-canonical structures in genome.

  12. Quantitative cognitive-test characterization of reconnectable implantable fiber-optic neurointerfaces for optogenetic neurostimulation.

    Science.gov (United States)

    Fedotov, I V; Ivashkina, O I; Pochechuev, M S; Roshchina, M A; Toropova, K A; Fedotov, A B; Anokhin, K V; Zheltikov, A M

    2017-02-23

    Cognitive tests on representative groups of freely behaving transgenic mice are shown to enable a quantitative characterization of reconnectable implantable fiber-optic neurointerfaces for optogenetic neurostimulation. A systematic analysis of such tests provides a robust quantitative measure for the cognitive effects induced by fiber-optic neurostimulation, validating the performance of fiber-optic neurointerfaces for long-term optogenetic brain stimulations and showing no statistically significant artifacts in the behavior of transgenic mice due to interface implantation. © 2017 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  13. Optimization Efficiency of Monte Carlo Simulation Tool for Evanescent Wave Spectroscopy Fiber-Optic Probe

    Directory of Open Access Journals (Sweden)

    Daniel Khankin

    2012-01-01

    Full Text Available In a previous work, we described the simulation tool (FOPS 3D (Khankin et al., 2001 which can simulate the full three-dimensional geometrical structure of a fiber and the propagation of a light beam sent through it. In this paper we are focusing on three major points: the first concerns the improvements made with respect to the simulation tool and the second, optimizations implemented with respect to the calculations' efficiency. Finally, the major research improvement from our previous works is the simulation results of the optimal absorbance value, as a function of bending angle for a given uncladded part diameter, that are presented; it is suggested that fiber-bending may improve the efficiency of recording the relevant measurements. This is the third iteration of the FOPS development process (Mann et al., 2009 which was significantly optimized by decreasing memory usage and increasing CPU utilization.

  14. Direct immobilization of DNA probes on non-modified plastics by UV irradiation and integration in microfluidic devices for rapid bioassay

    DEFF Research Database (Denmark)

    Yi, Sun; Perch-Nielsen, Ivan R.; Dufva, Martin

    2012-01-01

    and parallel identification of Avian Influenza Virus by DNA hybridization. The one-step, cost-effective DNA-linking method on non-modified polymers significantly simplifies microarray fabrication procedures and permits great flexibility to plastic material selection, thus making it convenient to integrate...

  15. Design of high-capacity fiber-optic transport systems

    Science.gov (United States)

    Liao, Zhi Ming

    2001-08-01

    We study the design of fiber-optic transport systems and the behavior of fiber amplifiers/lasers with the aim of achieving higher capacities with larger amplifier spacing. Solitons are natural candidates for transmitting short pulses for high-capacity fiber-optic networks because of its innate ability to use two of fiber's main defects, fiber dispersion and fiber nonlinearity to balance each other. In order for solitons to retain its dynamic nature, amplifiers must be placed periodically to restore powers to compensate for fiber loss. Variational analysis is used to study the long-term stability of a periodical- amplifier system. A new regime of operation is identified which allows the use of a much longer amplifier spacing. If optical fibers are the blood vessels of an optical communication system, then the optical amplifier based on erbium-doped fiber is the heart. Optical communication systems can avoid the use of costly electrical regenerators to maintain system performance by being able to optically amplify the weakened signals. The length of amplifier spacing is largely determined by the gain excursion experienced by the solitons. We propose, model, and demonstrate a distributed erbium-doped fiber amplifier which can drastically reduce the amount of gain excursion experienced by the solitons, therefore allowing a much longer amplifier spacing and superior stability. Dispersion management techniques have become extremely valuable tools in the design of fiber-optic communication systems. We have studied in depth the advantage of different arnplification schemes (lumped and distributed) for various dispersion compensation techniques. We measure the system performance through the Q factor to evaluate the added advantage of effective noise figure and smaller gain excursion. An erbium-doped fiber laser has been constructed and characterized in an effort to develop a test bed to study transmission systems. The presence of mode-partition noise in an erbium

  16. Study on Interaction between Apigenin and Herring Sperm DNA by Acridine Orange as a Fluorescence Probe%吖啶橙为荧光探针研究芹菜素与DNA的相互作用

    Institute of Scientific and Technical Information of China (English)

    尚永辉; 李华; 孙家娟; 刘彬

    2011-01-01

    在pH值为7.40的Tris-HC1缓冲溶液中,采用吸收光谱法、荧光光谱法以及粘度法研究了芹菜素(Ap)与鲱鱼精DNA(fsDNA)的相互作用.研究表明,Ap与fsDNA相互作用生成了结合比nAp:nDNA=2:1的复合物,温度300 K和310 K的结合常数Kb分别为1.068×104 L·m01-1和1.137×104 L·mol-1;300 K温度下Ap与DNA相互作用的△rHm为1.899×103 J·mol-1,△rSm为83.475 J·mo1-1·K-1,△rGm为-2.306×104 J·mo1-1,表明两者的结合过程为熵驱动反应.粘度测定结果进一步确定实验条件下Ap与fsDNA的作用方式为插入模式.%The interaction of apigenin with herring sperm DNA was studied with acridine orange as a fluorescence probe. The fluorescence spectra indicated that a kind of compound of apigenin and herring sperm DNA was formed at pH = 7. 40. The binary compound ratio was napigenin:nDNA = 2:1; the binding constants of apigenin with herring sperm DNA compound were 1. 068×104 L·mol-1 (300 K) and 1. 137× 104L·mol-1 (310 K) respectively. The thermodynamic parameters of the interaction were calculated as follows:△rHm = l. 899× 103 J·mol-1,△rSm = 83. 475 J·mol-1·K-1, △rGm =-2. 306×104 J·mol-1 at the 300 K. The interaction of apigenin with DNA was also studied through method of viscosity. The results confirmed that the intercalation model was the major mode of the interaction between apigenin and herring sperm DNA, and the binding of apigenin with herring sperm DNA was an entropy-driven reaction.

  17. Tunable nanowire nonlinear optical probe

    Energy Technology Data Exchange (ETDEWEB)

    Nakayama, Yuri; Pauzauskie, Peter J.; Radenovic, Aleksandra; Onorato, Robert M.; Saykally, Richard J.; Liphardt, Jan; Yang, Peidong

    2008-02-18

    One crucial challenge for subwavelength optics has been thedevelopment of a tunable source of coherent laser radiation for use inthe physical, information, and biological sciences that is stable at roomtemperature and physiological conditions. Current advanced near-fieldimaging techniques using fiber-optic scattering probes1,2 have alreadyachieved spatial resolution down to the 20-nm range. Recently reportedfar-field approaches for optical microscopy, including stimulatedemission depletion (STED)3, structured illumination4, and photoactivatedlocalization microscopy (PALM)5, have also enabled impressive,theoretically-unlimited spatial resolution of fluorescent biomolecularcomplexes. Previous work with laser tweezers6-8 has suggested the promiseof using optical traps to create novel spatial probes and sensors.Inorganic nanowires have diameters substantially below the wavelength ofvisible light and have unique electronic and optical properties9,10 thatmake them prime candidates for subwavelength laser and imagingtechnology. Here we report the development of an electrode-free,continuously-tunable coherent visible light source compatible withphysiological environments, from individual potassium niobate (KNbO3)nanowires. These wires exhibit efficient second harmonic generation(SHG), and act as frequency converters, allowing the local synthesis of awide range of colors via sum and difference frequency generation (SFG,DFG). We use this tunable nanometric light source to implement a novelform of subwavelength microscopy, in which an infrared (IR) laser is usedto optically trap and scan a nanowire over a sample, suggesting a widerange of potential applications in physics, chemistry, materials science,and biology.

  18. Fiber-Optic Vibration Sensor Based on Multimode Fiber

    Directory of Open Access Journals (Sweden)

    I. Lujo

    2008-06-01

    Full Text Available The purpose of this paper is to present a fiberoptic vibration sensor based on the monitoring of the mode distribution in a multimode optical fiber. Detection of vibrations and their parameters is possible through observation of the output speckle pattern from the multimode optical fiber. A working experimental model has been built in which all used components are widely available and cheap: a CCD camera (a simple web-cam, a multimode laser in visible range as a light source, a length of multimode optical fiber, and a computer for signal processing. Measurements have shown good agreement with the actual frequency of vibrations, and promising results were achieved with the amplitude measurements although they require some adaptation of the experimental model. Proposed sensor is cheap and lightweight and therefore presents an interesting alternative for monitoring large smart structures.

  19. Giant endobronchial hamartoma resected by fiberoptic bronchoscopy electrosurgical snaring

    Directory of Open Access Journals (Sweden)

    Cavallari Vittorio

    2011-08-01

    Full Text Available Abstract Less than 1% of lung neoplasms are represented by benign tumors. Among these, hamartomas are the most common with an incidence between 0.025% and 0.32%. In relation to the localization, hamartomas are divided into intraparenchymal and endobronchial. Clinical manifestation of an endobronchial hamartoma (EH results from tracheobronchial obstruction or bleeding. Usually, EH localizes in large diameter bronchus. Endoscopic removal is usually recommended. Bronchotomy or parenchimal resection through thoracotomy should be reserved only for cases where the hamatoma cannot be approached through endoscopy, or when irreversible lung functional impairment occurred after prolonged airflow obstruction. Generally, when endoscopic approach is used, this is through rigid bronchoscopy, laser photocoagulation or mechanical resection. Here we present a giant EH occasionally diagnosed and treated by fiberoptic bronchoscopy electrosurgical snaring.

  20. Fiber-optic gyro location of dome azimuth

    Science.gov (United States)

    Kuehne, John W.

    2016-07-01

    The 2.1-m Otto Struve Telescope, world's second largest in 1939, today has modern motion control and superb tracking, yet the 19-m-diameter Art Deco dome has resisted many attempts to record its azimuth electronically. Demonstrated in January 2016, a small tactical-grade fiber-optic gyro located anywhere on the rotating structure, aided by a few fiducial points to zero gyro drift, adequately locates the azimuth. The cost of a gyro is practically independent of dome size, offering an economical solution for large domes that cannot be easily encoded with conventional systems. The 100-Hz sampling is capable of revealing anomalies in the rotation rate, valuable for preventive maintenance on any dome. I describe software methods and time series analysis to integrate angular velocity to dome azimuth; transformation of telescope hour angle and declination into required dome azimuth, using a formula that accounts for a cross-axis mount inside an offset dome; and test results.

  1. Transoral tracheal intubation of rodents using a fiberoptic laryngoscope.

    Science.gov (United States)

    Costa, D L; Lehmann, J R; Harold, W M; Drew, R T

    1986-06-01

    A fiberoptic laryngoscope which allows direct visualization of the deep pharynx and epiglottis has been developed for transoral tracheal intubation of small laboratory mammals. The device has been employed in the intubation and instillation of a variety of substances into the lungs of rats, and with minor modification, has had similar application in mice, hamsters, and guinea pigs. The simplicity and ease of handling of the laryngoscope permits one person to intubate large numbers of enflurane anesthetized animals either on an open counter top or in a glove-box, as may be required for administration of carcinogenic materials. Instillation of 7Be-labeled carbon particles into the lungs of mice, hamsters, rats, and guinea pigs resulted in reasonably consistent interlobal distribution of particles for each test animal species with minimal tracheal deposition. However, actual lung tissue doses of carbon exhibited some species dependence.

  2. Miniaturized fiber-optic Michelson-type interferometric sensors

    Science.gov (United States)

    Murphy, Kent A.; Miller, William V., III; Tran, Tuan A.; Vengsarkar, Ashish M.; Claus, Richard O.

    1991-01-01

    A novel, miniaturized Michelson-type fiber-optic interferometric sensor that is relatively insensitive to temperature drifts is presented. A fused-biconical tapered coupler is cleaved immediately after the coupled length and polished down to the region of the fused cladding, but short of the interaction region. The end of one core is selectively coated with a reflective surface and is used as the reference arm; the other core serves as the sensing arm. The detection of surface acoustic waves, microdisplacements, and magnetic fields is reported. The sensor is shown to be highly stable in comparison to a classic homodyne, uncompensated Michelson interferometer, and signal-to-noise ratios of 65 dB have been obtained.

  3. Broadband excitation and collection in fiber-optic nonlinear endomicroscopy

    Science.gov (United States)

    Prakash Ghimire, Navin; Bao, Hongchun; Gu, Min

    2013-08-01

    Broadband excitation and collection in a fiber-optic nonlinear endomicroscope are realized by using a single hollow-core double-clad photonic crystal fiber and a gradient index lens. Femtosecond pulses with central wavelengths in the range of 750-850 nm can be directly delivered through the core of the fiber for nonlinear excitation without pre-chirping. A gradient index lens with numerical aperture 0.8 designed to operate over the near-infrared wavelength range is used for focusing the laser beam from the fiber for nonlinear excitation and for collecting the fluorescent signal from the sample. This compact system is suitable to perform nonlinear imaging of multiple fluorophors in the wavelength range of 750-850 nm.

  4. New composite fiber-optic overhead ground wire

    Science.gov (United States)

    Nishiyama, S.; Kitayama, Y.; Ona, A.; Shimada, S.; Kikuta, T.

    1986-11-01

    A composite fiber-optic overhead ground wire (OPGW) has already been used commercially. Most of the electric power companies have the plans to establish new telecommunication networks by means of OPGW, and it is greatly needed to develop OPGW which has a maximum number of fibers for specific ground wire size with low transmission loss, easy handling and higher reliability. We have developed New OPGW, which satisfies these demands. It has the following features: (1) 18 fibers within 75 sq mm ground wire. (2) The fiber is a pure silica core and a fluorine-doped cladding single mode fiber with the average transmission loss of 0.4 dB/km at 1.3 micron. (3) Six fibers are stranded into a compact bunch and three bunches are housed in the spiral grooves of an aluminum spacer.

  5. Fiber-Optic Shape Sensing for Intelligent Solar Sail Deployment Project

    Data.gov (United States)

    National Aeronautics and Space Administration — Luna Innovations proposes to develop a distributed fiber-optic shape sensor to provide a control system for the deployment of ultra-lightweight inflatable support...

  6. High-Frequency Flush Mounted Miniature LOX Fiber-Optic Pressure Sensor II Project

    Data.gov (United States)

    National Aeronautics and Space Administration — Luna Innovations has teamed with the University of Alabama, Huntsville, to develop a miniature flush-mounted fiber-optic pressure sensor that will allow accurate,...

  7. High-Frequency Flush Mounted Miniature LOX Fiber-Optic Pressure Sensor Project

    Data.gov (United States)

    National Aeronautics and Space Administration — Luna Innovations is teaming with the University of Alabama, Huntsville, to develop a miniature flush-mounted fiber-optic pressure sensor that will allow accurate,...

  8. FIBER-OPTIC BIOSENSOR FOR DIRECT DETERMINATION OF ORGANOPHOSPHATE NERVE AGENTS. (R823663)

    Science.gov (United States)

    A fiber-optic enzyme biosensor for the direct measurement of organophosphate nerveagents was developed. The basic element of this biosensor is organophosphorus hydrolaseimmobilized on a nylon membrane and attached to the common end of a bifurcated optical fiberbundle....

  9. Frequency-Shifted Interferometry — A Versatile Fiber-Optic Sensing Technique

    Directory of Open Access Journals (Sweden)

    Fei Ye

    2014-06-01

    Full Text Available Fiber-optic sensing is a field that is developing at a fast pace. Novel fiber-optic sensor designs and sensing principles constantly open doors for new opportunities. In this paper, we review a fiber-optic sensing technique developed in our research group called frequency-shifted interferometry (FSI. This technique uses a continuous-wave light source, an optical frequency shifter, and a slow detector. We discuss the operation principles of several FSI implementations and show their applications in fiber length and dispersion measurement, locating weak reflections along a fiber link, fiber-optic sensor multiplexing, and high-sensitivity cavity ring-down measurement. Detailed analysis of FSI system parameters is also presented.

  10. Construction of genome-wide physical BAC contigs using mapped cDNA as probes: Toward an integrated BAC library resource for genome sequencing and analysis. Annual report, July 1995--January 1997

    Energy Technology Data Exchange (ETDEWEB)

    Mitchell, S.C.; Bocskai, D.; Cao, Y. [and others

    1997-12-31

    The goal of human genome project is to characterize and sequence entire genomes of human and several model organisms, thus providing complete sets of information on the entire structure of transcribed, regulatory and other functional regions for these organisms. In the past years, a number of useful genetic and physical markers on human and mouse genomes have been made available along with the advent of BAC library resources for these organisms. The advances in technology and resource development made it feasible to efficiently construct genome-wide physical BAC contigs for human and other genomes. Currently, over 30,000 mapped STSs and 27,000 mapped Unigenes are available for human genome mapping. ESTs and cDNAs are excellent resources for building contig maps for two reasons. Firstly, they exist in two alternative forms--as both sequence information for PCR primer pairs, and cDoreen genomic libraries efficiently for large number of DNA probes by combining over 100 cDNA probes in each hybridization. Second, the linkage and order of genes are rather conserved among human, mouse and other model organisms. Therefore, gene markers have advantages over random anonymous STSs in building maps for comparative genomic studies.

  11. Dispersed dynamics of solvation in G-quadruplex DNA: comparison of dynamic Stokes shifts of probes in parallel and antiparallel quadruplex structures

    Science.gov (United States)

    Kiran Singh, Moirangthem; Shweta, Him; Sen, Sobhan

    2016-09-01

    G-quadruplex DNA (GqDNA) structures play an important role in many specific cellular functions and are promising anti-tumor targets for small molecules (ligands). Here, we measured the dynamic Stokes shift of a ligand (Hoechst) bound to parallel c-Myc (mPu22) GqDNA over five decades of time from 100 fs to 10 ns, and compared it with the previously reported dynamics of DAPI bound to antiparallel human telomeric (hTelo22) GqDNA (Pal et al 2015 J. Phys. Chem. Lett. 6 1754). Stokes shift data from fluorescence up-conversion and time-correlated single photon counting experiments was combined to cover the broad dynamic range. The results show that the solvation dynamics of Hoechst in parallel mPu22 GqDNA follow a power law relaxation, added to fast 2 ps exponential relaxation, from 100 fs to 10 ns, with only a subtle difference of power law exponents in the two ligand-GqDNA systems (0.06 in Hoechst-mPu22 compared to 0.16 in DAPI-hTelo22). We measured steady-state fluorescence spectra and time-resolved anisotropy decays which confirm the tight binding of Hoechst to parallel mPu22 with a binding constant of ~1  ×  105 M-1. The molecular docking of Hoechst in parallel GqDNA followed by a 50 ns molecular dynamics (MD) simulation on a Hoechst-GqDNA complex reveals that Hoechst binds to one of the outer G-tetrads by end-stacking near G13 and G4, which is different from the binding site of DAPI inside a groove of antiparallel hTelo22 GqDNA. Reconciling previous experimental and simulation results, we assign the 2 ps component to the hydration dynamics of only weakly perturbed water near mPu22 and the power law relaxation to the coupled motion of water and DNA (i.e. DNA backbone, unpaired bases and loops connecting G-tetrads) which come near the Hoechst inside parallel GqDNA.

  12. Probe Storage

    NARCIS (Netherlands)

    Gemelli, Marcellino; Abelmann, Leon; Engelen, Johan B.C.; Khatib, Mohammed G.; Koelmans, Wabe W.; Zaboronski, Olog; Campardo, Giovanni; Tiziani, Federico; Laculo, Massimo

    2011-01-01

    This chapter gives an overview of probe-based data storage research over the last three decades, encompassing all aspects of a probe recording system. Following the division found in all mechanically addressed storage systems, the different subsystems (media, read/write heads, positioning, data chan

  13. Cultural probes

    DEFF Research Database (Denmark)

    Madsen, Jacob Østergaard

    2016-01-01

    The aim of this study was thus to explore cultural probes (Gaver, Boucher et al. 2004), as a possible methodical approach, supporting knowledge production on situated and contextual aspects of occupation.......The aim of this study was thus to explore cultural probes (Gaver, Boucher et al. 2004), as a possible methodical approach, supporting knowledge production on situated and contextual aspects of occupation....

  14. A two-step stimulus-response cell-SELEX method to generate a DNA aptamer to recognize inflamed human aortic endothelial cells as a potential in vivo molecular probe for atherosclerosis plaque detection.

    Science.gov (United States)

    Ji, Kaili; Lim, Wee Siang; Li, Sam Fong Yau; Bhakoo, Kishore

    2013-08-01

    Aptamers are single-stranded oligonucleotides that are capable of binding wide classes of targets with high affinity and specificity. Their unique three-dimensional structures present numerous possibilities for recognizing virtually any class of target molecules, making them a promising alternative to antibodies used as molecular probes in biomedical analysis and clinical diagnosis. In recent years, cell-systematic evolution of ligands by exponential enrichment (SELEX) has been used extensively to select aptamers for various cell targets. However, aptamers that have evolved from cell-SELEX to distinguish the "stimulus-response cell" have not previously been reported. Moreover, a number of cumbersome and time-consuming steps involved in conventional cell-SELEX reduce the efficiency and efficacy of the aptamer selection. Here, we report a "two-step" methodology of cell-SELEX that successfully selected DNA aptamers specifically against "inflamed" endothelial cells. This has been termed as stimulus-response cell-SELEX (SRC-SELEX). The SRC-SELEX enables the selection of aptamers to distinguish the cells activated by stimulus of healthy cells or cells isolated from diseased tissue. We report a promising aptamer, N55, selected by SRC-SELEX, which can bind specifically to inflamed endothelial cells both in cell culture and atherosclerotic plaque tissue. This aptamer probe was demonstrated as a potential molecular probe for magnetic resonance imaging to target inflamed endothelial cells and atherosclerotic plaque detection.

  15. Design of a fiber-optic transmitter for microwave analog transmission with high phase stability

    Science.gov (United States)

    Logan, R. T., Jr.; Lutes, G. F.; Primas, L. E.; Maleki, L.

    1990-01-01

    The principal considerations in the design of fiber-optic transmitters for highly phase-stable radio frequency and microwave analog transmission are discussed. Criteria for a fiber-optic transmitter design with improved amplitude and phase-noise performance are developed through consideration of factors affecting the phase noise, including low-frequency laser-bias supply noise, the magnitude and proximity of external reflections into the laser, and temperature excursions of the laser-transmitter package.

  16. Development of a novel polymeric fiber-optic magnetostrictive metal detector.

    Science.gov (United States)

    Hua, Wei-Shu; Hooks, Joshua Rosenberg; Wu, Wen-Jong; Wang, Wei-Chih

    2010-01-01

    The purpose this paper is the development a novel polymeric fiber-optic magnetostrictive metal detector, using a fiber-optic Mach-Zehnder interferometer and polymeric magnetostrictive material. Metal detection is based on the strain-induced optical path length change steming from the ferromagnetic material introduced in the magnetic field. Varied optical phase shifts resulted largely from different metal objects. In this paper, the preliminary results on the different metal material detection will be discussed.

  17. Fiber-optic fluorometer for microscale mapping of photosynthetic pigments in microbial communities

    DEFF Research Database (Denmark)

    Thar, Roland Matthias; Kühl, Michael; Holst, Gerhard

    2001-01-01

    Microscale fluorescence measurements were performed in photosynthetic biofilms at a spatial resolution of 100 to 200 µm with a new fiber-optic fluorometer which allowed four different excitation and emission wavelengths and was configured for measuring phycobiliproteins, chlorophylls, and bacteri......Microscale fluorescence measurements were performed in photosynthetic biofilms at a spatial resolution of 100 to 200 µm with a new fiber-optic fluorometer which allowed four different excitation and emission wavelengths and was configured for measuring phycobiliproteins, chlorophylls...

  18. A Fiber-Optical Intrusion Alarm System Based on Quasi-Distributed Fiber Bragg Grating Sensors

    Institute of Scientific and Technical Information of China (English)

    Qi Jiang; Yun-Jiang Rao; De-Hong Zeng

    2008-01-01

    A fiber-optical intrusion alarm system based on quasi-distributed fiber Bragg grating (FBG) sensors is demonstrated in this paper. The algorithms of empirical mode decomposition (EMD) and wavelet packet characteristic entropy are adopted to determine the intrusion location. The intrusion alarm software based on the Labview is developed, and it is also proved by the experiments. The results show that such a fiber-optical intrusion alarm system can offer the automatic intrusion alarm in real-time.

  19. Fiber-optic fluorometer for microscale mapping of photosynthetic pigments in microbial communities

    DEFF Research Database (Denmark)

    Thar, Roland Matthias; Kühl, Michael; Holst, Gerhard

    2001-01-01

    Microscale fluorescence measurements were performed in photosynthetic biofilms at a spatial resolution of 100 to 200 µm with a new fiber-optic fluorometer which allowed four different excitation and emission wavelengths and was configured for measuring phycobiliproteins, chlorophylls, and bacteri......Microscale fluorescence measurements were performed in photosynthetic biofilms at a spatial resolution of 100 to 200 µm with a new fiber-optic fluorometer which allowed four different excitation and emission wavelengths and was configured for measuring phycobiliproteins, chlorophylls...

  20. Hairpin DNA probe with 5'-TCC/CCC-3' overhangs for the creation of silver nanoclusters and miRNA assay.

    Science.gov (United States)

    Xia, Xiaodong; Hao, Yuanqiang; Hu, Shengqiang; Wang, Jianxiu

    2014-01-15

    A facile strategy for the assay of target miRNA using fluorescent silver nanoclusters (AgNCs) has been described. Due to the preferable interaction between cytosine residues and Ag(+), a short cytosine-rich oligonucleotide (ODN) with only six bases 5'-TCCCCC-3' served as an efficient scaffold for the creation of the AgNCs. The AgNCs displayed a bright red emission when excited at 545nm. Such ODN base-stabilized AgNCs have been exploited for miRNA sensing. Overhangs of TCC at the 5' end (5'-TCC) and CCC at the 3' end (CCC-3') (denoted as 5'-TCC/CCC-3') appended to the hairpin ODN probe which also contains recognition sequences for target miRNA were included. Interestingly, the AgNCs/hairpin ODN probe showed similar spectral properties as that templated by 5'-TCCCCC-3'. The formation of the hairpin ODN probe/miRNA duplex separated the 5'-TCC/CCC-3' overhangs, thus disturbing the optical property or structure of the AgNCs. As a result, fluorescence quenching of the AgNCs/hairpin ODN probe was obtained, which allows for facile determination of target miRNA. The proposed method is simple and cost-effective, holding great promise for clinical applications.