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Sample records for 9g dna chip

  1. From DNA biosensors to gene chips

    PubMed Central

    Wang, Joseph

    2000-01-01

    Wide-scale DNA testing requires the development of small, fast and easy-to-use devices. This article describes the preparation, operation and applications of biosensors and gene chips, which provide fast, sensitive and selective detection of DNA hybridization. Various new strategies for DNA biosensors and gene chips are examined, along with recent trends and future directions. The integration of hybridization detection schemes with the sample preparation process in a ‘Lab-on-a-Chip’ format is also covered. While the use of DNA biosensors and gene chips is at an early stage, such devices are expected to have an enormous effect on future DNA diagnostics. PMID:10931914

  2. Microarrays Made Simple: "DNA Chips" Paper Activity

    ERIC Educational Resources Information Center

    Barnard, Betsy

    2006-01-01

    DNA microarray technology is revolutionizing biological science. DNA microarrays (also called DNA chips) allow simultaneous screening of many genes for changes in expression between different cells. Now researchers can obtain information about genes in days or weeks that used to take months or years. The paper activity described in this article…

  3. Microarrays (DNA Chips) for the Classroom Laboratory

    ERIC Educational Resources Information Center

    Barnard, Betsy; Sussman, Michael; BonDurant, Sandra Splinter; Nienhuis, James; Krysan, Patrick

    2006-01-01

    We have developed and optimized the necessary laboratory materials to make DNA microarray technology accessible to all high school students at a fraction of both cost and data size. The primary component is a DNA chip/array that students "print" by hand and then analyze using research tools that have been adapted for classroom use. The primary…

  4. Ultrasensitive DNA chip: gene expression profile analysis without RNA amplification.

    PubMed

    Nagino, Kunihisa; Nomura, Osamu; Takii, Yuki; Myomoto, Akira; Ichikawa, Makiko; Nakamura, Fumio; Higasa, Masashi; Akiyama, Hideo; Nobumasa, Hitoshi; Shiojima, Satoshi; Tsujimoto, Gozoh

    2006-04-01

    We have developed a new DNA chip whose substrate has a unique minute columnar array structure made of plastic. The DNA chip exhibits ultrahigh sensitivity, up to 100-fold higher than that of reference DNA chips, which makes it possible to monitor gene expression profiles even with very small amounts of RNA (0.1-0.01 microg of total RNA) without amplification. Differential expression ratios obtained with the new DNA chip were validated against those obtained with quantitative real-time PCR assays. This novel microarray technology would be a powerful tool for monitoring gene expression profiles, especially for clinical diagnosis.

  5. Assembly of highly aligned DNA strands onto Si chips.

    PubMed

    Zhang, Jianming; Ma, Yufeng; Stachura, Sylwia; He, Huixin

    2005-04-26

    This paper reports a robust and efficient approach to assemble highly aligned DNA strands onto Si chips. The method combines advantages from molecular combing and microcontact printing to realize controlling both the density and direction of DNA strands on the Si chip. In addition, it also can be utilized to prepare stretched DNA structures on solid surfaces. Compared to approaches that use molecular combing directly on silanated surfaces, the stretched single-chain DNA structures are straighter. Furthermore, by exploiting the hydrophobic property of the intrinsic poly(dimethylsiloxane) stamp, this study also describes a simple way to produce straight bundled DNA arrays on Si and other substrates.

  6. Microarrays/DNA Chips for the Detection of Waterborne Pathogens.

    PubMed

    Vale, Filipa F

    2016-01-01

    DNA microarrays are useful for the simultaneous detection of microorganisms in water samples. Specific probes targeting waterborne pathogens are selected with bioinformatics tools, synthesized and spotted onto a DNA array. Here, the construction of a DNA chip for waterborne pathogen detection is described, including the processes of probe in silico selection, synthesis, validation, and data analysis. PMID:27460375

  7. Genetic screening with the DNA chip: a new Pandora's box?

    PubMed Central

    Henn, W

    1999-01-01

    The ethically controversial option of genetic population screening used to be restricted to a small number of rather rare diseases by methodological limitations which are now about to be overcome. With the new technology of DNA microarrays ("DNA chip"), emerging from the synthesis of microelectronics and molecular biology, methods are now at hand for the development of mass screening programmes for a wide spectrum of genetic traits. Thus, the DNA chip may be the key technology for a refined preventive medicine as well as a new dimension of eugenics. The forthcoming introduction of the DNA chip technology into medical practice urgently requires an internationally consistent framework of ethical standards and legal limitations if we do not want it to become a new Pandora's box. PMID:10226928

  8. Development of a protein microarray using sequence-specific DNA binding domain on DNA chip surface

    SciTech Connect

    Choi, Yoo Seong; Pack, Seung Pil; Yoo, Young Je . E-mail: yjyoo@snu.ac.kr

    2005-04-22

    A protein microarray based on DNA microarray platform was developed to identify protein-protein interactions in vitro. The conventional DNA chip surface by 156-bp PCR product was prepared for a substrate of protein microarray. High-affinity sequence-specific DNA binding domain, GAL4 DNA binding domain, was introduced to the protein microarray as fusion partner of a target model protein, enhanced green fluorescent protein. The target protein was oriented immobilized directly on the DNA chip surface. Finally, monoclonal antibody of the target protein was used to identify the immobilized protein on the surface. This study shows that the conventional DNA chip can be used to make a protein microarray directly, and this novel protein microarray can be applicable as a tool for identifying protein-protein interactions.

  9. DNA chips with conjugated polyelectrolytes in resonance energy transfer mode.

    PubMed

    Wigenius, Jens A; Magnusson, Karin; Björk, Per; Andersson, Olof; Inganäs, Olle

    2010-03-01

    We show how to use well-defined conjugated polyelectrolytes (CPEs) combined with surface energy patterning to fabricate DNA chips utilizing fluorescence signal amplification. Cholesterol-modified DNA strands in complex with a CPE are adsorbed to a surface energy pattern, formed by printing with soft elastomer stamps. Hybridization of the surface bound DNA strands with a short complementary strand from solution is monitored using both fluorescence microscopy and imaging surface plasmon resonance. The CPEs act as antennas, enhancing resonance energy transfer to the dye-labeled DNA when complementary hybridization of the double strand occurs.

  10. On-chip concentration of bacteria using a 3D dielectrophoretic chip and subsequent laser-based DNA extraction in the same chip

    NASA Astrophysics Data System (ADS)

    Cho, Yoon-Kyoung; Kim, Tae-hyeong; Lee, Jeong-Gun

    2010-06-01

    We report the on-chip concentration of bacteria using a dielectrophoretic (DEP) chip with 3D electrodes and subsequent laser-based DNA extraction in the same chip. The DEP chip has a set of interdigitated Au post electrodes with 50 µm height to generate a network of non-uniform electric fields for the efficient trapping by DEP. The metal post array was fabricated by photolithography and subsequent Ni and Au electroplating. Three model bacteria samples (Escherichia coli, Staphylococcus epidermidis, Streptococcus mutans) were tested and over 80-fold concentrations were achieved within 2 min. Subsequently, on-chip DNA extraction from the concentrated bacteria in the 3D DEP chip was performed by laser irradiation using the laser-irradiated magnetic bead system (LIMBS) in the same chip. The extracted DNA was analyzed with silicon chip-based real-time polymerase chain reaction (PCR). The total process of on-chip bacteria concentration and the subsequent DNA extraction can be completed within 10 min including the manual operation time.

  11. Identification of hepatotoxin-producing cyanobacteria by DNA-chip.

    PubMed

    Rantala, Anne; Rizzi, Ermanno; Castiglioni, Bianca; de Bellis, Gianluca; Sivonen, Kaarina

    2008-03-01

    We developed a new tool to detect and identify hepatotoxin-producing cyanobacteria of the genera Anabaena, Microcystis, Planktothrix, Nostoc and Nodularia. Genus-specific probe pairs were designed for the detection of the microcystin (mcyE) and nodularin synthetase genes (ndaF) of these five genera to be used with a DNA-chip. The method couples a ligation detection reaction, in which the polymerase chain reaction (PCR)-amplified mcyE/ndaF genes are recognized by the probe pairs, with a hybridization on a universal microarray. All the probe pairs specifically detected the corresponding mcyE/ndaF gene sequences when DNA from the microcystin- or nodularin-producing cyanobacterial strains were used as template in the PCR. Furthermore, the strict specificity of detection enabled identification of the potential hepatotoxin producers. Detection of the genes was very sensitive; only 1-5 fmol of the PCR product were needed to produce signal intensities that exceeded the set background threshold level. The genus-specific probe pairs also reliably detected potential microcystin producers in DNA extracted from six lake and four brackish water samples. In lake samples, the same microcystin producers were identified with quantitative real-time PCR analysis. The specificity, sensitivity and ability of the DNA-chip in simultaneously detecting all the main hepatotoxin producers make this method suitable for high-throughput analysis and monitoring of environmental samples.

  12. Avatar DNA Nanohybrid System in Chip-on-a-Phone

    NASA Astrophysics Data System (ADS)

    Park, Dae-Hwan; Han, Chang Jo; Shul, Yong-Gun; Choy, Jin-Ho

    2014-05-01

    Long admired for informational role and recognition function in multidisciplinary science, DNA nanohybrids have been emerging as ideal materials for molecular nanotechnology and genetic information code. Here, we designed an optical machine-readable DNA icon on microarray, Avatar DNA, for automatic identification and data capture such as Quick Response and ColorZip codes. Avatar icon is made of telepathic DNA-DNA hybrids inscribed on chips, which can be identified by camera of smartphone with application software. Information encoded in base-sequences can be accessed by connecting an off-line icon to an on-line web-server network to provide message, index, or URL from database library. Avatar DNA is then converged with nano-bio-info-cogno science: each building block stands for inorganic nanosheets, nucleotides, digits, and pixels. This convergence could address item-level identification that strengthens supply-chain security for drug counterfeits. It can, therefore, provide molecular-level vision through mobile network to coordinate and integrate data management channels for visual detection and recording.

  13. Avatar DNA Nanohybrid System in Chip-on-a-Phone

    PubMed Central

    Park, Dae-Hwan; Han, Chang Jo; Shul, Yong-Gun; Choy, Jin-Ho

    2014-01-01

    Long admired for informational role and recognition function in multidisciplinary science, DNA nanohybrids have been emerging as ideal materials for molecular nanotechnology and genetic information code. Here, we designed an optical machine-readable DNA icon on microarray, Avatar DNA, for automatic identification and data capture such as Quick Response and ColorZip codes. Avatar icon is made of telepathic DNA-DNA hybrids inscribed on chips, which can be identified by camera of smartphone with application software. Information encoded in base-sequences can be accessed by connecting an off-line icon to an on-line web-server network to provide message, index, or URL from database library. Avatar DNA is then converged with nano-bio-info-cogno science: each building block stands for inorganic nanosheets, nucleotides, digits, and pixels. This convergence could address item-level identification that strengthens supply-chain security for drug counterfeits. It can, therefore, provide molecular-level vision through mobile network to coordinate and integrate data management channels for visual detection and recording. PMID:24824876

  14. Avatar DNA nanohybrid system in chip-on-a-phone.

    PubMed

    Park, Dae-Hwan; Han, Chang Jo; Shul, Yong-Gun; Choy, Jin-Ho

    2014-05-14

    Long admired for informational role and recognition function in multidisciplinary science, DNA nanohybrids have been emerging as ideal materials for molecular nanotechnology and genetic information code. Here, we designed an optical machine-readable DNA icon on microarray, Avatar DNA, for automatic identification and data capture such as Quick Response and ColorZip codes. Avatar icon is made of telepathic DNA-DNA hybrids inscribed on chips, which can be identified by camera of smartphone with application software. Information encoded in base-sequences can be accessed by connecting an off-line icon to an on-line web-server network to provide message, index, or URL from database library. Avatar DNA is then converged with nano-bio-info-cogno science: each building block stands for inorganic nanosheets, nucleotides, digits, and pixels. This convergence could address item-level identification that strengthens supply-chain security for drug counterfeits. It can, therefore, provide molecular-level vision through mobile network to coordinate and integrate data management channels for visual detection and recording.

  15. Re-use of commercial microfluidics chips for DNA, RNA, and protein electrophoresis.

    PubMed

    Nguyen, Thi; Kwak, Sukyoung; Karpowicz, Steven J

    2014-11-01

    Microfluidics chip technology is a powerful and convenient alternative to agarose gels and PAGE, but costs can be high due to certain chips being non-reusable. Here we describe a method to regenerate, re-use, and store Agilent DNA, RNA, and protein electrophoresis chips designed for use in the Bioanalyzer 2100. By washing the sample wells and displacing the old gel matrix with new gel-dye mix, we have run samples on the same chip up to ten times with negligible loss of signal quality. Chips whose wells were loaded with buffer or water were stored successfully for one week before re-use.

  16. Microfluidic chip for stacking, separation and extraction of multiple DNA fragments.

    PubMed

    Wu, Ruige; Seah, Y P; Wang, Zhiping

    2016-03-11

    A disposable integrated microfluidic device was developed for rapid sample stacking, separation and extraction of multiple DNA fragments from a relatively large amount of sample. Isotachophoresis hyphenated gel electrophoresis (ITP-GE) was used to pre-concentrate and separate DNA fragments, followed by extraction of pure DNA fragments with electroelution on-chip. DNA fragments of 200bp, 500bp and 1kbp were successfully separated and collected in the extraction chamber within 25min. The extraction efficiency obtained from the chip was 49.9%, 52.1% and 53.7% for 200bp, 500bp and 1kbp DNA fragments, respectively. The extracted DNA fragments exhibited compatibility with downstream enzymatic reactions, for example PCR. The chip was also used to extract DNA fragments with specific size range from sheared genomic DNA and demonstrated similar performance to that using traditional gel cutting method. The whole assay can finish in 32min, 6 times faster than traditional method.

  17. Numerical modeling of DNA-chip hybridization with chaotic advection

    PubMed Central

    Raynal, Florence; Beuf, Aurélien; Carrière, Philippe

    2013-01-01

    We present numerical simulations of DNA-chip hybridization, both in the “static” and “dynamical” cases. In the static case, transport of free targets is limited by molecular diffusion; in the dynamical case, an efficient mixing is achieved by chaotic advection, with a periodic protocol using pumps in a rectangular chamber. This protocol has been shown to achieve rapid and homogeneous mixing. We suppose in our model that all free targets are identical; the chip has different spots on which the probes are fixed, also all identical, and complementary to the targets. The reaction model is an infinite sink potential of width dh, i.e., a target is captured as soon as it comes close enough to a probe, at a distance lower than dh. Our results prove that mixing with chaotic advection enables much more rapid hybridization than the static case. We show and explain why the potential width dh does not play an important role in the final results, and we discuss the role of molecular diffusion. We also recover realistic reaction rates in the static case. PMID:24404027

  18. Development of an Automated DNA Detection System Using an Electrochemical DNA Chip Technology

    NASA Astrophysics Data System (ADS)

    Hongo, Sadato; Okada, Jun; Hashimoto, Koji; Tsuji, Koichi; Nikaido, Masaru; Gemma, Nobuhiro

    A new compact automated DNA detection system Genelyzer™ has been developed. After injecting a sample solution into a cassette with a built-in electrochemical DNA chip, processes from hybridization reaction to detection and analysis are all operated fully automatically. In order to detect a sample DNA, electrical currents from electrodes due to an oxidization reaction of electrochemically active intercalator molecules bound to hybridized DNAs are detected. The intercalator is supplied as a reagent solution by a fluid supply unit of the system. The feasibility test proved that the simultaneous typing of six single nucleotide polymorphisms (SNPs) associated with a rheumatoid arthritis (RA) was carried out within two hours and that all the results were consistent with those by conventional typing methods. It is expected that this system opens a new way to a DNA testing such as a test for infectious diseases, a personalized medicine, a food inspection, a forensic application and any other applications.

  19. A Novel Self-Assembling DNA Nano Chip for Rapid Detection of Human Papillomavirus Genes

    PubMed Central

    Li, Xin; Li, Yanbo; Hong, Li

    2016-01-01

    Rapid detection of tumor-associated DNA such as Human Papillomavirus (HPV) has important clinical value for the early screening of tumors. By attaching oligonucleotides or cDNA onto the chip surface, DNA chip technology provides a rapid method to analyze gene expression. However, challenges remain regarding increasing probe density and improving detection time. To address these challenges, we proposed a DNA chip that was self-assembled from single stranded DNA in combination with high probe density and a rapid detection method. Over 200 probes could be attached to the surface of this 100-nm diameter DNA chip. For detection, the chips were adsorbed onto a mica surface and then incubated for ten minutes with HPV-DNA; the results were directly observable using atomic force microscopy (AFM). This bottom-up fabricated DNA nano chip combined with high probe density and direct AFM detection at the single molecule level will likely have numerous potential clinical applications for gene screening and the early diagnosis of cancer. PMID:27706184

  20. Addressable microfluidic polymer chip for DNA-directed immobilization of oligonucleotide-tagged compounds.

    PubMed

    Schröder, Hendrik; Hoffmann, Linda; Müller, Joachim; Alhorn, Petra; Fleger, Markus; Neyer, Andreas; Niemeyer, Christof M

    2009-07-01

    A microfluidic polymer chip for the self-assembly of DNA conjugates through DNA-directed immobilization is developed. The chip is fabricated from two parts, one of which contains a microfluidic channel produced from poly(dimethylsiloxane) (PDMS) by replica-casting technique using a mold prepared by photolithographic techniques. The microfluidic part is sealed by covalent bonding with a chemically activated glass slide containing a DNA oligonucleotide microarray. The dimension of the PDMS-glass microfluidic chip is equivalent to standard microscope slides (76 x 26 mm(2)). The DNA microarray surface inside the microfluidic channels is configured through conventional spotting, and the resulting DNA patches can be conveniently addressed with compounds containing complementary DNA tags. To demonstrate the utility of the addressable surface within the microfluidic channel, DNA-directed immobilization (DDI) of DNA-modified gold nanoparticles (AuNPs) and DNA-conjugates of the enzymes glucose oxidase (GOx) and horseradish peroxidase (HRP) are carried out. DDI of AuNPs is used to demonstrate site selectivity and reversibility of the surface-modification process. In the case of the DNA-enzyme conjugates, the patterned assembly of the two enzymes allows the establishment and investigation of the coupled reaction of GOx and HRP, with particular emphasis on surface coverage and lateral flow rates. The results demonstrate that this addressable chip is well suited for the generation of fluidically coupled multi-enzyme microreactors.

  1. Radio Frequency Identification Sensor Chips with Anticollision Algorithm for Simultaneous Detection of Multiple DNA Targets

    NASA Astrophysics Data System (ADS)

    Yazawa, Yoshiaki; Oonishi, Tadashi; Watanabe, Kazuki; Nemoto, Ryo; Shiratori, Akiko

    2010-04-01

    A newly developed DNA measurement method for multiple single nucleotide polymorphism (SNP) typing using a radio-frequency identification (RFID) sensor chip was demonstrated. The RFID sensor chip monolithically integrates a sensor, amplifier, analog-to-digital converter (ADC), and a passive wireless communication interface for receiving commands and transmitting data on a 2.5×2.5 mm2 silicon chip. For the simultaneous multitarget measurement, anticollision control and peak-power suppression are essential. To assign a unique identification number (UID) for the identification of multiple sensor chips, a reproducible random number generator circuit (RRG) was designed and installed on the chip. Peak-power consumption was reduced to 1018 µW by a clock gating of functional circuit blocks. Multiple SNP typing was carried out by simultaneously operating five RFID sensor chips (four with photosensors and one with a temperature sensor). The target DNA was captured on the sensor chips, and SNPs were detected by observing bioluminescence. Finally, the observed data were wirelessly transmitted to the reader.

  2. Continuous cell electroporation for efficient DNA and siRNA delivery based on laminar microfluidic chips.

    PubMed

    Wei, Zewen; Li, Zhihong

    2014-01-01

    Electroporation is a high-efficiency and low-toxicity physical gene transfer method. Traditional electroporation is limited to only low volume cell samples. Here we present a continuous cell electroporation method based on commonly used microfluidic chip fabrication technology. Using easily fabricated PDMS microfluidic chip, syringe pumps, and pulse generator, we show efficient delivery of both DNA and siRNA into different cell lines. We describe the protocol of chip fabrication, apparatus setup, and cell electroporation assay. Typically, the fabrication of the devices takes 1 or 2 days and the continuous electroporation assay takes 1 h.

  3. DNA-affinity-purified chip (DAP-chip) method to determine gene targets for bacterial two component regulatory systems.

    PubMed

    Rajeev, Lara; Luning, Eric G; Mukhopadhyay, Aindrila

    2014-07-21

    In vivo methods such as ChIP-chip are well-established techniques used to determine global gene targets for transcription factors. However, they are of limited use in exploring bacterial two component regulatory systems with uncharacterized activation conditions. Such systems regulate transcription only when activated in the presence of unique signals. Since these signals are often unknown, the in vitro microarray based method described in this video article can be used to determine gene targets and binding sites for response regulators. This DNA-affinity-purified-chip method may be used for any purified regulator in any organism with a sequenced genome. The protocol involves allowing the purified tagged protein to bind to sheared genomic DNA and then affinity purifying the protein-bound DNA, followed by fluorescent labeling of the DNA and hybridization to a custom tiling array. Preceding steps that may be used to optimize the assay for specific regulators are also described. The peaks generated by the array data analysis are used to predict binding site motifs, which are then experimentally validated. The motif predictions can be further used to determine gene targets of orthologous response regulators in closely related species. We demonstrate the applicability of this method by determining the gene targets and binding site motifs and thus predicting the function for a sigma54-dependent response regulator DVU3023 in the environmental bacterium Desulfovibrio vulgaris Hildenborough.

  4. Automatic on-chip RNA-DNA hybridization assay with integrated phase change microvalves

    NASA Astrophysics Data System (ADS)

    Weng, Xuan; Jiang, Hai; Wang, Junsheng; Chen, Shu; Cao, Honghe; Li, Dongqing

    2012-07-01

    An RNA-DNA hybridization assay microfluidic chip integrated with electrothermally actuated phase change microvalves for detecting pathogenic bacteria is presented in this paper. In order to realize the sequential loading and washing processes required in such an assay, gravity-based pressure-driven flow and phase-change microvalves were used in the microfluidic chip. Paraffin wax was used as the phase change material in the valves and thin film heaters were used to electrothermally actuate microvalves. Light absorption measured by a photodetector to determine the concentrations of the samples. The automatic control of the complete assay was implemented by a self-coded LabVIEW program. To examine the performance of this chip, Salmonella was used as a sample pathogen. Significantly, reduction in reagent/sample consumption (up to 20 folds) was achieved by this on-chip assay, compared with using the commercial test kit following the same protocol in conventional labs. The experimental results show that the quantitative detection can be obtained in approximately 26 min, and the detection limit is as low as 103 CFU ml-1. This RNA-DNA hybridization assay microfluidic chip shows an excellent potential in the development of a portable device for point-of-testing applications.

  5. Series DNA Amplification Using the Continuous-Flow Polymerase Chain Reaction Chip

    NASA Astrophysics Data System (ADS)

    Joung, Seung-Ryong; Kang, Chi Jung; Kim, Yong-Sang

    2008-02-01

    We proposed a continuous-flow polymerase chain reaction (PCR) chip that can be used for series DNA amplification. The continuous-flow PCR chip has several advantages such as fast thermal cycling, series of amplifications, cost-effective fabrication, portability, and fluorescence detection. The continuous-flow PCR chip is composed of two parts namely poly(dimethylsiloxane) (PDMS) microchannel for sample injection and indium-tin-oxide (ITO) heater/glass chip for thermal cycling. The fabricated microchannel width and depth are 250 and 200 µm, respectively. Also, the total working length of the PDMS microchannel is 1340 mm which is equivalent for 20 cycles of amplification. A 2:2:3 microchannel length ratio for three different temperature zones namely denaturation, annealing, and extension was assigned, respectively. Upon the operation of the fabricated continuous-flow PCR chip, the amplification of plasmid DNA pKS-GFP with 720 base pairs and PG-noswsi with 300 base pairs were found successfully with a total reaction time of 15 min.

  6. Disposable on-chip microfluidic system for buccal cell lysis, DNA purification, and polymerase chain reaction.

    PubMed

    Cho, Woong; Maeng, Joon-Ho; Ahn, Yoomin; Hwang, Seung Yong

    2013-09-01

    This paper reports the development of a disposable, integrated biochip for DNA sample preparation and PCR. The hybrid biochip (25 × 45 mm) is composed of a disposable PDMS layer with a microchannel chamber and reusable glass substrate integrated with a microheater and thermal microsensor. Lysis, purification, and PCR can be performed sequentially on this microfluidic device. Cell lysis is achieved by heat and purification is performed by mechanical filtration. Passive check valves are integrated to enable sample preparation and PCR in a fixed sequence. Reactor temperature is needed to lysis and PCR reaction is controlled within ±1°C by PID controller of LabVIEW software. Buccal epithelial cell lysis, DNA purification, and SY158 gene PCR amplification were successfully performed on this novel chip. Our experiments confirm that the entire process, except the off-chip gel electrophoresis, requires only approximately 1 h for completion. This disposable microfluidic chip for sample preparation and PCR can be easily united with other technologies to realize a fully integrated DNA chip.

  7. Molecular Basis of 9G4 B Cell Autoreactivity in Human Systemic Lupus Erythematosus

    PubMed Central

    Richardson, Christopher; Chida, Asiya Seema; Adlowitz, Diana; Silver, Lin; Fox, Erin; Jenks, Scott A.; Palmer, Elise; Wang, Youliang; Heimburg-Molinaro, Jamie; Li, Quan-Zhen; Mohan, Chandra; Cummings, Richard; Tipton, Christopher

    2013-01-01

    9G4+ IgG Abs expand in systemic lupus erythematosus (SLE) in a disease-specific fashion and react with different lupus Ags including B cell Ags and apoptotic cells. Their shared use of VH4-34 represents a unique system to understand the molecular basis of lupus autoreactivity. In this study, a large panel of recombinant 9G4+ mAbs from single naive and memory cells was generated and tested against B cells, apoptotic cells, and other Ags. Mutagenesis eliminated the framework-1 hydrophobic patch (HP) responsible for the 9G4 idiotype. The expression of the HP in unselected VH4-34 cells was assessed by deep sequencing. We found that 9G4 Abs recognize several Ags following two distinct structural patterns. B cell binding is dependent on the HP, whereas anti-nuclear Abs, apoptotic cells, and dsDNA binding are HP independent and correlate with positively charged H chain third CDR. The majority of mutated VH4-34 memory cells retain the HP, thereby suggesting selection by Ags that require this germline structure. Our findings show that the germline-encoded HP is compulsory for the anti–B cell reactivity largely associated with 9G4 Abs in SLE but is not required for reactivity against apoptotic cells, dsDNA, chromatin, anti-nuclear Abs, or cardiolipin. Given that the lupus memory compartment contains a majority of HP+ VH4-34 cells but decreased B cell reactivity, additional HP-dependent Ags must participate in the selection of this compartment. This study represents the first analysis, to our knowledge, of VH-restricted autoreactive B cells specifically expanded in SLE and provides the foundation to understand the antigenic forces at play in this disease. PMID:24108696

  8. Fenton fragmentation for faster electrophoretic on chip purification of amplifiable genomic DNA.

    PubMed

    Hakenberg, S; Hügle, M; Meyer, P; Behrmann, O; Dame, G; Urban, G A

    2015-05-15

    With a rapid and simple actuation protocol electrophoretic nucleic acid extraction is easy automatable, requires no moving parts, is easy to miniaturize and furthermore possesses a size dependent cut-off filter adjustable by the pore size of the hydrogel. However electrophoretic nucleic acid extraction from bacteria has so far been applied mainly for short RNA targets. One of the reasons is that electrophoretic processing of unfragmented genomic DNA strands is time-consuming, because of the length. Here DNA fragmentation would accelerate extraction and isolation. We introduce on-chip lysis and non-enzymatic DNA cleavage directly followed by a purifying step for receiving amplifiable DNA fragments from bacteria in less than 25 min. In contrast to restriction enzymes the Fenton reaction is known to cleave DNA without nucleotide specificity. The reaction mix contains iron(II) EDTA, sodium ascorbate, hydrogen peroxide and lysozyme. The degree of fragmentation can be adjusted by the concentration of reagents. The results enable electrophoretic extraction methods to unspecifically process long genomic DNA in a short time frame, e.g. for pathogen detection in a lab-on-a-chip format.

  9. Fenton fragmentation for faster electrophoretic on chip purification of amplifiable genomic DNA.

    PubMed

    Hakenberg, S; Hügle, M; Meyer, P; Behrmann, O; Dame, G; Urban, G A

    2015-05-15

    With a rapid and simple actuation protocol electrophoretic nucleic acid extraction is easy automatable, requires no moving parts, is easy to miniaturize and furthermore possesses a size dependent cut-off filter adjustable by the pore size of the hydrogel. However electrophoretic nucleic acid extraction from bacteria has so far been applied mainly for short RNA targets. One of the reasons is that electrophoretic processing of unfragmented genomic DNA strands is time-consuming, because of the length. Here DNA fragmentation would accelerate extraction and isolation. We introduce on-chip lysis and non-enzymatic DNA cleavage directly followed by a purifying step for receiving amplifiable DNA fragments from bacteria in less than 25 min. In contrast to restriction enzymes the Fenton reaction is known to cleave DNA without nucleotide specificity. The reaction mix contains iron(II) EDTA, sodium ascorbate, hydrogen peroxide and lysozyme. The degree of fragmentation can be adjusted by the concentration of reagents. The results enable electrophoretic extraction methods to unspecifically process long genomic DNA in a short time frame, e.g. for pathogen detection in a lab-on-a-chip format. PMID:24970713

  10. On-chip magnetic bead-based DNA melting curve analysis using a magnetoresistive sensor

    NASA Astrophysics Data System (ADS)

    Rizzi, Giovanni; Østerberg, Frederik W.; Henriksen, Anders D.; Dufva, Martin; Hansen, Mikkel F.

    2015-04-01

    We present real-time measurements of DNA melting curves in a chip-based system that detects the amount of surface-bound magnetic beads using magnetoresistive magnetic field sensors. The sensors detect the difference between the amount of beads bound to the top and bottom sensor branches of the differential sensor geometry. The sensor surfaces are functionalized with wild type (WT) and mutant type (MT) capture probes, differing by a single base insertion (a single nucleotide polymorphism, SNP). Complementary biotinylated targets in suspension couple streptavidin magnetic beads to the sensor surface. The beads are magnetized by the field arising from the bias current passed through the sensors. We demonstrate the first on-chip measurements of the melting of DNA hybrids upon a ramping of the temperature. This overcomes the limitation of using a single washing condition at constant temperature. Moreover, we demonstrate that a single sensor bridge can be used to genotype a SNP.

  11. Magnetoresistive DNA chips based on ac field focusing of magnetic labels

    NASA Astrophysics Data System (ADS)

    Ferreira, H. A.; Cardoso, F. A.; Ferreira, R.; Cardoso, S.; Freitas, P. P.

    2006-04-01

    A study was made on the sensitivity of a magnetoresistive DNA-chip platform being developed for cystic fibrosis diagnostics. The chip, comprised of an array of 2.5×80 μm2 U-shaped spin-valve sensors integrated within current line structures for magnetic label manipulation, enabled the detection at 30 Hz of 250 nm magnetic nanoparticles from 100 pM down to the pM range (or a target DNA concentration of 500 pM). It was observed that the sensor response increased linearly with label concentration. Noise spectra obtained for these sensors showed a thermal noise of 10-17 V2/Hz with a 1/f knee at 50 kHz at a 1 mA sense current, showing that lower detection limits are possible.

  12. DNA-library assembly programmed by on-demand nano-liter droplets from a custom microfluidic chip

    PubMed Central

    Tangen, Uwe; Minero, Gabriel Antonio S.; Sharma, Abhishek; Wagler, Patrick F.; Cohen, Rafael; Raz, Ofir; Marx, Tzipy; Ben-Yehezkel, Tuval; McCaskill, John S.

    2015-01-01

    Nanoscale synthetic biology can benefit from programmable nanoliter-scale processing of DNA in microfluidic chips if they are interfaced effectively to biochemical arrays such as microwell plates. Whereas active microvalve chips require complex fabrication and operation, we show here how a passive and readily fabricated microchip can be employed for customizable nanoliter scale pipetting and reaction control involving DNA. This recently developed passive microfluidic device, supporting nanoliter scale combinatorial droplet generation and mixing, is here used to generate a DNA test library with one member per droplet exported to addressed locations on microwell plates. Standard DNA assembly techniques, such as Gibson assembly, compatible with isothermal on-chip operation, are employed and checked using off-chip PCR and assembly PCR. The control of output droplet sequences and mixing performance was verified using dyes and fluorescently labeled DNA solutions, both on-chip and in external capillary channels. Gel electrophoresis of products and DNA sequencing were employed to further verify controlled combination and functional enzymatic assembly. The scalability of the results to larger DNA libraries is also addressed by combinatorial input expansion using sequential injection plugs from a multiwell plate. Hence, the paper establishes a proof of principle of the production of functional combinatorial mixtures at the nanoliter scale for one sequence per well DNA libraries. PMID:26221198

  13. Analysis of DNA-chip and antigen-chip data: studies of cancer, stem cells and autoimmune diseases

    NASA Astrophysics Data System (ADS)

    Domany, Eytan

    2005-07-01

    Biology has undergone a revolution during the past decade. Deciphering the human genome has opened new horizons, among which the advent of DNA microarrays has been perhaps the most significant. These miniature measuring devices report the levels at which tens of thousands of genes are expressed in a collection of cells of interest (such as tissue from a tumor). I describe here briefly this technology and present an example of how analysis of data obtained from such high throughput experiments provides insights of possible clinical and therapeutic relevance for Acute Lymphoblastic Leukemia. Next, I describe how gene expression data is used to deduce a new design principle, " Just In Case", used by stem cells. Finally I briefly review a different novel technology, of antigen chips, which provide a fingerprint of a subject's immune system and may become a predictive clinical tool. The work reviewed here was done in collaboration with numerous colleagues and students.

  14. Electrochemical chip-based genomagnetic assay for detection of high-risk human papillomavirus DNA.

    PubMed

    Bartosik, Martin; Durikova, Helena; Vojtesek, Borivoj; Anton, Milan; Jandakova, Eva; Hrstka, Roman

    2016-09-15

    Cervical cancer, being the fourth leading cause of cancer death in women worldwide, predominantly originates from a persistent infection with a high-risk human papillomavirus (HPV). Detection of DNA sequences from these high-risk strains, mostly HPV-16 and HPV-18, represents promising strategy for early screening, which would help to identify women with higher risk of cervical cancer. In developing countries, inadequate screening options lead to disproportionately high mortality rates, making a fast and inexpensive detection schemes highly important. Electrochemical sensors and assays offer an alternative to current methods of detection. We developed an electrochemical-chip based assay, in which target HPV DNA is captured via magnetic bead-modified DNA probes, followed by an antidigoxigenin-peroxidase detection system at screen-printed carbon electrode chips, enabling parallel measurements of eight samples simultaneously. We show sensitive detection in attomoles of HPV DNA, selective discrimination between HPV-16 and HPV-18 and good reproducibility. Most importantly, we show application of the assay into both cancer cell lines and cervical smears from patients. The electrochemical results correlated well with standard methods, making this assay potentially applicable in clinical practice.

  15. On-chip DNA preconcentration in different media conductivities by electrodeless dielectrophoresis

    PubMed Central

    Li, Shunbo; Ye, Ziran; Hui, Yu Sanna; Gao, Yibo; Jiang, Yusheng; Wen, Weijia

    2015-01-01

    Electrodeless dielectrophoresis is the best choice to achieve preconcentration of nanoparticles and biomolecules due to its simple, robust, and easy implementation. We designed a simple chip with microchannels and nano-slits in between and then studied the trapping of DNA in high conductive medium and low conductive medium, corresponding to positive and negative dielectrophoresis (DEP), respectively. It is very important to investigate the trapping in media with different conductivities since one always has to deal with the sample solutions with different conductivities. The trapping process was analyzed by the fluorescent intensity changes. The results showed that DNA could be trapped at the nano-slit in both high and low conductive media in a lower electric field strength (10 V/cm) compared to the existing methods. This is a significant improvement to suppress the Joule heating effect in DEP related experiments. Our work may give insight to researchers for DNA trapping by a simple and low cost device in the Lab-on-a-Chip system. PMID:26487901

  16. On-chip DNA preconcentration in different media conductivities by electrodeless dielectrophoresis.

    PubMed

    Li, Shunbo; Ye, Ziran; Hui, Yu Sanna; Gao, Yibo; Jiang, Yusheng; Wen, Weijia

    2015-09-01

    Electrodeless dielectrophoresis is the best choice to achieve preconcentration of nanoparticles and biomolecules due to its simple, robust, and easy implementation. We designed a simple chip with microchannels and nano-slits in between and then studied the trapping of DNA in high conductive medium and low conductive medium, corresponding to positive and negative dielectrophoresis (DEP), respectively. It is very important to investigate the trapping in media with different conductivities since one always has to deal with the sample solutions with different conductivities. The trapping process was analyzed by the fluorescent intensity changes. The results showed that DNA could be trapped at the nano-slit in both high and low conductive media in a lower electric field strength (10 V/cm) compared to the existing methods. This is a significant improvement to suppress the Joule heating effect in DEP related experiments. Our work may give insight to researchers for DNA trapping by a simple and low cost device in the Lab-on-a-Chip system. PMID:26487901

  17. High-speed DNA genotyping using microfabricated capillary array electrophoresis chips

    SciTech Connect

    Woolley, A.T.; Sensabaugh, G.F.; Mathies, R.A.

    1997-06-01

    Capillary array electrophoresis (CAE) chips have been designed and fabricated with the capacity to rapidly (<160 s) analyze 12 different samples in parallel. Detection of all lanes with 0.3 s temporal resolution was achieved using a laser-excited confocal-fluorescence scanner. The operation and capabilities of these CAE microdevices were first determined by performing electrophoretic separations of pBR322 MspI DNA samples. Genotyping of HLA-H, a candidate gene for the diagnosis of hereditary hemochromatosis, was then performed to demonstrate the rapid analysis of biologically relevant samples. Two-color multiplex fluorescence detection of HLA-H genotypes was accomplished by prelabeling the standard pBR322 MspI DNA ladder with a red emitting bisintercalation dye (butyl TOTIN) and on-column labeling of the HLA-H DNA with thiazole orange. This work establishes the feasibility of using CAE chips for high-speed, high-throughput genotyping. 44 refs., 7 figs.

  18. Novel developments for improved detection of specific mRNAs by DNA chips.

    PubMed

    Pioch, Daniel; Schweder, Thomas; Jürgen, Britta

    2008-10-01

    Microarrays have revolutionized gene expression analysis as they allow for highly parallel monitoring of mRNA levels of thousands of genes in a single experiment. Since their introduction some 15 years ago, substantial progress has been achieved with regard to, e.g., faster or more sensitive analyses. In this review, interesting new approaches for a more sensitive detection of specific mRNAs will be highlighted. Particularly, the potential of electrical DNA chip formats that allow for faster mRNA analyses will be discussed.

  19. Miniaturized devices towards an integrated lab-on-a-chip platform for DNA diagnostics

    NASA Astrophysics Data System (ADS)

    Kaprou, G.; Papadakis, G.; Kokkoris, G.; Papadopoulos, V.; Kefala, I.; Papageorgiou, D.; Gizeli, E.; Tserepi, A.

    2015-06-01

    Microfluidics is an emerging technology enabling the development of Lab-on-a-chip (LOC) systems for clinical diagnostics, drug discovery and screening, food safety and environmental analysis. LOC systems integrate and scale down one or several laboratory functions on a single chip of a few mm2 to cm2 in size, and account for many advantages on biochemical analyses, such as low sample and reagent consumption, low cost, reduced analysis time, portability and point-of-need compatibility. Currently, available nucleic acid diagnostic tests take advantage of Polymerase Chain Reaction (PCR) that allows exponential amplification of portions of nucleic acid sequences that can be used as indicators for the identification of various diseases. Here, we present a comparison between static chamber and continuous flow miniaturized PCR devices, in terms of energy consumption for devices fabricated on the same material stack, with identical sample volume and channel dimensions. The comparison is implemented by a computational study coupling heat transfer in both solid and fluid, mass conservation of species, and joule heating. Based on the conclusions of this study, we develop low-cost and fast DNA amplification devices for both PCR and isothermal amplification, and we implement them in the detection of mutations related to breast cancer. The devices are fabricated by mass production amenable technologies on printed circuit board (PCB) substrates, where copper facilitates the incorporation of on-chip microheaters, defining the thermal zones necessary for PCR or isothermal amplification methods.

  20. In silico evaluation of a novel DNA chip based fingerprinting technology for viral identification.

    PubMed

    Méndez-Tenorio, Alfonso; Flores-Cortés, Perla; Guerra-Trejo, Armando; Jaimes-Díaz, Hueman; Reyes-Rosales, Emma; Maldonado-Rodríguez, Arcadio; Espinosa-Lara, Mercedes; Maldonado-Rodríguez, Rogelio; Kenneth, Loren Beattie

    2006-01-01

    The identification of microorganisms by whole genome DNA fingerprinting was tested "in silico". 94 HPV genome sequences were submitted to virtual hybridization analysis on a DNA chip with 342 probes. This Universal Fingerprinting Chip (UFC) constitutes a representative set of probes of all the possible 8-mer sequences having at least two internal and non contiguous sequence differences between all them. A virtual hybridization analysis was performed in order to find the fingerprinting pattern that represents the signals produced for the hybridization of the probes allowing at most a single mismatch. All the fingerprints for each virus were compared against each other in order to obtain all the pairwise distances measures. A match-extension strategy was applied to identify only the shared signals corresponding to the hybridization of the probes with homologous sequences between two HPV genomes. A phylogenetic tree was constructed from the fingerprint distances using the Neighbor-Joining algorithm implemented in the program Phylip 3.61. This tree was compared with that produced from the alignment of whole genome HPV sequences calculated with the program Clustal_X 1.83. The similarities between both trees are suggesting that the UFC-8 is able to discriminate accurately between viral genomes. A fingerprint comparative analysis suggests that the UFC-8 can differentiate between HPV types and sub-types. PMID:17578073

  1. [The development of reagents set in the format of DNA-chip for genetic typing of strains of Vibrio cholerae].

    PubMed

    Pudova, E A; Markelov, M L; Dedkov, V G; Tchekanova, T A; Sadjin, A I; Kirdiyashkina, N P; Bekova, M V; Deviyatkin, A A

    2014-05-01

    The necessity of development of methods of genic diagnostic of cholera is conditioned by continuation of the Seventh pandemic of cholera, taxonomic variability of strains of Vibrio cholerae involved into pandemic and also permanent danger of delivery of disease to the territory of the Russian Federation. The methods of genic diagnostic of cholera make it possible in a comparatively short time to maximally minutely characterize strains isolated from patients or their environment. The article presents information about working out reagents set for genetic typing of agents of cholera using DNA-chip. The makeup of DNA-chip included oligonucleotide probes making possible to differentiate strains of V. cholerae on serogroups and biovars and to determine their pathogenicity. The single DNA-chip makes it possible to genetically type up to 12 samples concurrently. At that, duration of analysis without accounting stage of DNA separation makes up to 5 hours. In the progress of work, 23 cholera and non-cholera strains were analyzed. The full compliance of DNA-chip typing results to previously known characteristics of strains. Hence, there is a reason to consider availability of further development of reagents set and possibility of its further application in laboratories of regional level and reference centers. PMID:25338464

  2. Nanofluidic laboratory-on-chip device for mapping of single molecule DNA extracted from single cells

    NASA Astrophysics Data System (ADS)

    Mahshid, Sara; Berard, Daniel; Sladek, Robert; Leslie, Sabrina; Reisner, Walter

    2014-03-01

    The aim of this project is to create a nanofluidic platform to provide comprehensive maps of single-cell genomes at 1 kbp resolution based on the direct analysis of single 1-10 Mbp extended DNA molecules extracted from individual cells on-chip. We have developed a nanodevice in which all biochemical processing of single cells (cell lysis, DNA purification and fragmentation) is performed in situ. The platform has the following three components: (1) a micro-cavity (50 ×20 micron in dimension) for trapping and biochemical processing of single cells; (2) post arrays (1 micron depth) for untangling the released genomic contents and (3) parallel nanochannel arrays (100 nm) for extension of ~ 1-10 Mbp DNA for high-throughput optical mapping. Moreover, we use ``Convex Lense-Induced Nanoconfinement'' (CLIC) technique for trapping of single cell and dragging DNA into nanochannels. The principle is that a convex lens is pushed down to deform a flexible coverslip lid above the aforesaid platform containing nano/micro patterns, creating a locally confined region that pins molecules in the embedded nano/micro features. CLIC is used to lower the device lid over a cell isolated in the microcavity with an adjustable gap for buffer exchange. The released DNA is untangled using 1 micron-deep post arrays and driven into nanochannel array where its genomic content is revealed. In particular, using CLIC we were able to successfully trap 20 micron lymphoblast cells inside microcavity and lyse the trapped cell to drive out DNA.

  3. Biodetection of DNA and proteins using enhanced UV absorption by structuration of the chip surface

    NASA Astrophysics Data System (ADS)

    Robin, K.; Reverchon, J. L.; Mugherli, L.; Fromant, M.; Benisty, H.

    2009-02-01

    DNA and protein absorption at 260 and 280 nm can be used to reveal theses species on a biochip UV image. A first study including the design and fabrication of UV reflective multilayer biochips designed for UV contrast enhancement (factor of 4.0) together with spectrally selective AlGaN detectors demonstrated the control of chip biological coating, or Antigen/Antibody complexation with fairly good signals for typical probe density of 4x1012 molecules/cm2. Detection of fractional monolayer molecular binding requires a higher contrast enhancement which can be obtained with structured chips. Grating structures enable, at resonance, a confinement of light at the biochip surface, and thus a large interaction between the biological molecule and the lightwave field. The highest sensitivity obtained with grating-based biochip usually concerns a resonance shift, in wavelength or diffraction angle. Diffraction efficiency is also affected by UV absorption, due to enhanced light-matter interaction, and this mechanism is equally able to produce biochip images in parallel. By adjusting grating parameters, we will see how a biochip that is highly sensitive to UV absorption at its surface can be obtained. Based on the Ewald construction and diffraction diagram, instrumental resolution and smarter experimental configurations are considered. Notably, in conjunction with the 2D UV-sensitive detectors recently developed in-house, we discuss the obtainment of large contrast and good signals in a diffraction order emerging around the sample normal.

  4. Rapid detection for primary screening of influenza A virus: microfluidic RT-PCR chip and electrochemical DNA sensor.

    PubMed

    Yamanaka, Keiichiro; Saito, Masato; Kondoh, Kenji; Hossain, Mohammad Mosharraf; Koketsu, Ritsuko; Sasaki, Tadahiro; Nagatani, Naoki; Ikuta, Kazuyoshi; Tamiya, Eiichi

    2011-05-21

    Rapid and definitive diagnosis is critical to the prevention of the spread of endemic human pathogenic viruses. Detection of variant specific genes by reverse transcription polymerase chain reaction (RT-PCR) has become a routine diagnostic test for accurate subtyping of RNA viruses, such as influenza. In this paper, we demonstrate the use of a continuous-flow polydimethylsiloxane (PDMS) microfluidic RT-PCR chip and disposable electrical printed (DEP) chips for rapid amplification and sensing of new influenza (AH1pdm) virus of swine-origin. The RT-PCR chip consisted of four zones: RT reaction zone, initial denaturation zone, thermal cycle zone for PCR (2-step PCR) and pressurizing-channel zone for preventing air-bubble formation. In order to measure electrochemical signals, methylene blue (MB), an electro-active DNA intercalator, was added to the RT-PCR mixture. The RT-PCR was completed within 15 min which was the total flow-through time from the inlet to the outlet, and the reduction signals from amplifications could be detected quickly on the DEP chip. The MB reduction current on the DEP chip with the amplicon significantly reduced compared to non-amplified controls. This microfluidic platform for rapid RT-PCR and the DEP chip for quick electrochemical sensing are suitable for integration, and have the potential to be a portable system for diagnostic tests.

  5. Baseball bats and chocolate chip cookies: the judicial treatment of DNA in the myriad genetics litigation.

    PubMed

    Binnie, Ian; Park-Thompson, Vanessa

    2015-06-01

    In June 2013, the U.S. Supreme Court rendered a controversial ruling that naturally occurring DNA segments are "products of nature" and therefore not patentable subject matter. At this intersection between science and law, in litigation of crucial importance to patients, science, and multibillion-dollar biotech enterprises, the appellate judges sidestepped genetics and engaged in a war of metaphors from diamonds to chocolate chip cookies. This case is not an outlier. Apprehensive judges and juries in both Canada and the United States find many convenient excuses to avoid coming to grips with the underlying science in patent cases. But this is simply not acceptable. Legal rulings must be, and must seem to be, well grounded, as a matter of both law and science. The legitimacy of court decisions in the eyes of the stakeholders and the broader public depends on it. PMID:25524722

  6. Baseball bats and chocolate chip cookies: the judicial treatment of DNA in the myriad genetics litigation.

    PubMed

    Binnie, Ian; Park-Thompson, Vanessa

    2014-12-18

    In June 2013, the U.S. Supreme Court rendered a controversial ruling that naturally occurring DNA segments are "products of nature" and therefore not patentable subject matter. At this intersection between science and law, in litigation of crucial importance to patients, science, and multibillion-dollar biotech enterprises, the appellate judges sidestepped genetics and engaged in a war of metaphors from diamonds to chocolate chip cookies. This case is not an outlier. Apprehensive judges and juries in both Canada and the United States find many convenient excuses to avoid coming to grips with the underlying science in patent cases. But this is simply not acceptable. Legal rulings must be, and must seem to be, well grounded, as a matter of both law and science. The legitimacy of court decisions in the eyes of the stakeholders and the broader public depends on it.

  7. Detection of pathogens using on-chip electrochemical analysis of PCR amplified DNA molecules

    NASA Astrophysics Data System (ADS)

    Hodko, Dalibor; Raymer, Lindsay; Herbst, Stephanie M.; Magnuson, James W.; Gaskin, David

    2001-05-01

    The sensitivity and speed of methods for the detection of microorganisms and/or cells need to be constantly improved to provide timely and accurate analysis in large number of important applications. Such applications range from detection of pathogens in drinking water, biological warfare agents, biomedical diagnostics and food industry. The trends toward miniaturization of sensors using microfluidic and nanofluidic on-chip devices will push current detection limits to lower concentrations than what is offered by the present analytical equipment and/or detection kids. Microfluidic devices have been used to perform DNA analysis, polymerase chain reaction analysis, capillary electrophoresis and hybridization to oligonucleotide probes. This paper describes a new approach for the detection of pathogens on contaminated surfaces, which will integrated sampling, concentration and detection of targeted microorganisms.

  8. Genomic Sequencing and Biological Characteristics of a Novel Escherichia Coli Bacteriophage 9g, a Putative Representative of a New Siphoviridae Genus

    PubMed Central

    Kulikov, Eugene E.; Golomidova, Alla K.; Letarova, Maria A.; Kostryukova, Elena S.; Zelenin, Alexandr S.; Prokhorov, Nikolai S.; Letarov, Andrey V.

    2014-01-01

    Bacteriophage 9g was isolated from horse feces using Escherichia coli C600 as a host strain. Phage 9g has a slightly elongated capsid 62 × 76 nm in diameter and a non-contractile tail about 185 nm long. The complete genome sequence of this bacteriophage consists of 56,703 bp encoding 70 predicted open reading frames. The closest relative of phage 9g is phage PhiJL001 infecting marine alpha-proteobacterium associated with Ircinia strobilina sponge, sharing with phage 9g 51% of amino acid identity in the main capsid protein sequence. The DNA of 9g is resistant to most restriction endonucleases tested, indicating the presence of hypermodified bases. The gene cluster encoding a biosynthesis pathway similar to biosynthesis of the unusual nucleoside queuosine was detected in the phage 9g genome. The genomic map organization is somewhat similar to the typical temperate phage gene layout but no integrase gene was detected. Phage 9g efficiently forms stable associations with its host that continues to produce the phage over multiple passages, but the phage can be easily eliminated via viricide treatment indicating that no true lysogens are formed. Since the sequence, genomic organization and biological properties of bacteriophage 9g are clearly distinct from other known Enterobacteriaceae phages, we propose to consider it as the representative of a novel genus of the Siphoviridae family. PMID:25533657

  9. Genomic sequencing and biological characteristics of a novel Escherichia coli bacteriophage 9g, a putative representative of a new Siphoviridae genus.

    PubMed

    Kulikov, Eugene E; Golomidova, Alla K; Letarova, Maria A; Kostryukova, Elena S; Zelenin, Alexandr S; Prokhorov, Nikolai S; Letarov, Andrey V

    2014-12-01

    Bacteriophage 9 g was isolated from horse feces using Escherichia coli C600 as a host strain. Phage 9 g has a slightly elongated capsid 62 × 76 nm in diameter and a non-contractile tail about 185 nm long. The complete genome sequence of this bacteriophage consists of 56,703 bp encoding 70 predicted open reading frames. The closest relative of phage 9 g is phage PhiJL001 infecting marine alpha-proteobacterium associated with Ircinia strobilina sponge, sharing with phage 9 g 51% of amino acid identity in the main capsid protein sequence. The DNA of 9 g is resistant to most restriction endonucleases tested, indicating the presence of hypermodified bases. The gene cluster encoding a biosynthesis pathway similar to biosynthesis of the unusual nucleoside queuosine was detected in the phage 9 g genome. The genomic map organization is somewhat similar to the typical temperate phage gene layout but no integrase gene was detected. Phage 9 g efficiently forms stable associations with its host that continues to produce the phage over multiple passages, but the phage can be easily eliminated via viricide treatment indicating that no true lysogens are formed. Since the sequence, genomic organization and biological properties of bacteriophage 9 g are clearly distinct from other known Enterobacteriaceae phages, we propose to consider it as the representative of a novel genus of the Siphoviridae family. PMID:25533657

  10. CometChip: a high-throughput 96-well platform for measuring DNA damage in microarrayed human cells.

    PubMed

    Ge, Jing; Prasongtanakij, Somsak; Wood, David K; Weingeist, David M; Fessler, Jessica; Navasummrit, Panida; Ruchirawat, Mathuros; Engelward, Bevin P

    2014-10-18

    DNA damaging agents can promote aging, disease and cancer and they are ubiquitous in the environment and produced within human cells as normal cellular metabolites. Ironically, at high doses DNA damaging agents are also used to treat cancer. The ability to quantify DNA damage responses is thus critical in the public health, pharmaceutical and clinical domains. Here, we describe a novel platform that exploits microfabrication techniques to pattern cells in a fixed microarray. The 'CometChip' is based upon the well-established single cell gel electrophoresis assay (a.k.a. the comet assay), which estimates the level of DNA damage by evaluating the extent of DNA migration through a matrix in an electrical field. The type of damage measured by this assay includes abasic sites, crosslinks, and strand breaks. Instead of being randomly dispersed in agarose in the traditional assay, cells are captured into an agarose microwell array by gravity. The platform also expands from the size of a standard microscope slide to a 96-well format, enabling parallel processing. Here we describe the protocols of using the chip to evaluate DNA damage caused by known genotoxic agents and the cellular repair response followed after exposure. Through the integration of biological and engineering principles, this method potentiates robust and sensitive measurements of DNA damage in human cells and provides the necessary throughput for genotoxicity testing, drug development, epidemiological studies and clinical assays.

  11. Semi-automated bacterial spore detection system with micro-fluidic chips for aerosol collection, spore treatment and ICAN DNA detection.

    PubMed

    Inami, Hisao; Tsuge, Kouichiro; Matsuzawa, Mitsuhiro; Sasaki, Yasuhiko; Togashi, Shigenori; Komano, Asuka; Seto, Yasuo

    2009-07-15

    A semi-automated bacterial spore detection system (BSDS) was developed to detect biological threat agents (e.g., Bacillus anthracis) on-site. The system comprised an aerosol sampler, micro-fluidic chip-A (for spore germination and cell lysis), micro-fluidic chip-B (for extraction and detection of genomic DNA) and an analyzer. An aerosol with bacterial spores was first collected in the collection chamber of chip-A with a velocity of 300 l/min, and the chip-A was taken off from the aerosol sampler and loaded into the analyzer. Reagents packaged in the chip-A were sequentially applied into the chamber. The genomic DNA extract from spore lyzate was manually transferred from chip-A to chip-B and loaded into the analyzer. Genomic DNA in chip-B was first trapped on a glass bead column, washed with various reagents, and eluted to the detection chamber by sequential auto-dispensing. Isothermal and chimeric primer-initiated amplification of nucleic acids (ICAN) with fluorescent measurement was adopted to amplify and detect target DNA. Bacillus subtilis was the stimulant of biological warfare agent in this experiment. Pretreatment conditions were optimized by examining bacterial target DNA recovery in the respective steps (aerosol collection, spore germination, cell lysis, and DNA extraction), by an off-chip experiment using a real-time polymerase chain reaction quantification method. Without the germination step, B. subtilis spores did not demonstrate amplification of target DNA. The detection of 10(4) spores was achieved within 2h throughout the micro-fluidic process. PMID:19450964

  12. Application of an electric DNA-chip for the expression analysis of bioprocess-relevant marker genes of Bacillus subtilis.

    PubMed

    Jürgen, Britta; Barken, Kim Bundvig; Tobisch, Steffen; Pioch, Daniel; Wümpelmann, Mogens; Hecker, Michael; Schweder, Thomas

    2005-11-01

    The knowledge of critical process-relevant genes can be used for an improved control of bioprocesses. So far bioprocess-relevant marker genes can be analyzed by established expression analysis methods only off-line. In this study, an alternative approach for a potential at-line monitoring of gene expression during bioprocesses is suggested. This approach is based on the measurement of specific mRNAs on an electric DNA-chip in connection with a magnetic bead-based sandwich hybridization. In order to allow an at-line measurement of specific mRNAs an improved method for a fast and partially automated isolation of high quality-RNA samples was developed. The expression analysis of the electric DNA-chip was compared with optical DNA micro arrays and the real time RT-PCR for three selected process-relevant genes of Bacillus subtilis. We demonstrate that the mRNA analysis by means of the electric DNA-chip gives similar results compared to the micro array analysis and the real time RT-PCR technique.

  13. PMA-PhyloChip DNA Microarray to Elucidate Viable Microbial Community Structure

    NASA Technical Reports Server (NTRS)

    Venkateswaran, Kasthuri J.; Stam, Christina N.; Andersen, Gary L.; DeSantis, Todd

    2011-01-01

    in the dark. Thereafter, the sample is exposed to visible light for five minutes, so that the DNA from dead cells will be cross-linked. Following this PMA treatment step, the sample is concentrated by centrifugation and washed (to remove excessive PMA) before DNA is extracted. The 16S rRNA gene fragments will be amplified by PCR to screen the total microbial community using PhyloChip DNA microarray analysis. This approach will detect only the viable microbial community since the PMA intercalated DNA from dead cells would be unavailable for PCR amplification. The total detection time including PCR reaction for low biomass samples will be a few hours. Numerous markets may use this technology. The food industry uses spore detection to validate new alternative food processing technologies, sterility, and quality. Pharmaceutical and medical equipment companies also detect spores as a marker for sterility. This system can be used for validating sterilization processes, water treatment systems, and in various public health and homeland security applications.

  14. Natural DNA variation at candidate loci is associated with potato chip color, tuber starch content, yield and starch yield.

    PubMed

    Li, Li; Paulo, Maria-João; Strahwald, Josef; Lübeck, Jens; Hofferbert, Hans-Reinhard; Tacke, Eckhart; Junghans, Holger; Wunder, Jörg; Draffehn, Astrid; van Eeuwijk, Fred; Gebhardt, Christiane

    2008-05-01

    Complex characters of plants such as starch and sugar content of seeds, fruits, tubers and roots are controlled by multiple genetic and environmental factors. Understanding their molecular basis will facilitate diagnosis and combination of superior alleles in crop improvement programs ("precision breeding"). Association genetics based on candidate genes is one approach toward this goal. Tetraploid potato varieties and breeding clones related by descent were evaluated for 2 years for chip quality before and after cold storage, tuber starch content, yield and starch yield. Chip quality is inversely correlated with tuber sugar content. A total of 36 loci on 11 potato chromosomes were evaluated for natural DNA variation in 243 individuals. These loci included microsatellites and genes coding for enzymes that function in carbohydrate metabolism or transport (candidate loci). The markers were used to analyze population structure and were tested for association with the tuber quality traits. Highly significant and robust associations of markers with 1-4 traits were identified. Most frequent were associations with chip quality and tuber starch content. Alleles increasing tuber starch content improved chip quality and vice versa. With two exceptions, the most significant and robust associations (q < 0.01) were observed with DNA variants in genes encoding enzymes that function in starch and sugar metabolism or transport. Comparing linkage and linkage disequilibrium between loci provided evidence for the existence of large haplotype blocks in the breeding materials analyzed.

  15. DNA mutation detection with chip-based temperature gradient capillary electrophoresis using a slantwise radiative heating system.

    PubMed

    Zhang, Hui-Dan; Zhou, Jing; Xu, Zhang-Run; Song, Jin; Dai, Jing; Fang, Jin; Fang, Zhao-Lun

    2007-09-01

    A simple and robust chip-based temperature gradient capillary electrophoresis (TGCE) system was developed for DNA mutation/single-nucleotide polymorphism (SNP) analysis using a radiative heating system. Reproducible, stable and uniform temperature gradients were established along a 3 cm length of the electrophoretic separation channel using a single thermostated aluminium heater plate. The heater was slightly slanted relative to the plane of the glass chip at 0.2-1.3 degrees by inserting thin spacers between the plate and chip at one end to produce differences in radiative heating that created the temperature gradient. On-chip TGCE analyses of 4 mutant DNA model samples amplified from plasmid templates, each containing a single base substitution, with a wide range of melting temperatures, showed that mutations were successfully detected under a wide temperature gradient of 10 degrees C and within a short gradient region of about 3 cm (3.3 degrees C cm(-1) gradient). The radiative heating system was able to establish stable spatial temperature gradients along short microfluidic separation channels using simple peripheral equipment and manipulation while ensuring good resolution for detecting a wide range of mutations. Effectiveness of the system was demonstrated by the successful detection of K-ras gene mutations in 6 colon cancer cell lines.

  16. Separation of large DNA molecules by size exclusion chromatography-based microchip with on-chip concentration structure

    NASA Astrophysics Data System (ADS)

    Azuma, Naoki; Itoh, Shintaro; Fukuzawa, Kenji; Zhang, Hedong

    2016-06-01

    The separation of DNA molecules according to their size represents a fundamental bioanalytical procedure. Here, we report the development of a chip-sized device, consisting of micrometer-sized fence structures fabricated in a microchannel, for the separation of large DNA molecules (over 10 kbp) based on the principle of size exclusion chromatography (SEC). In order to achieve separation, two approaches were utilized: first, the DNA samples were concentrated immediately prior to separation using nanoslit structures, with the aim of improving the resolution. Second, a theoretical model of SEC-based separation was established and applied in order to predict the optimal voltage range for separation. In this study, we achieved separation of λ DNA (48.5 kbp) and T4 DNA (166 kbp) using the present SEC-based microchip.

  17. Ultrasensitive Label-free Electronic Chip for DNA Analysis Using Carbon Nanotube Nanoelectrode Arrays

    NASA Technical Reports Server (NTRS)

    Li, Jun; Koehne, Jessica; Chen, Hua; Cassell, Alan; Ng, Hou Tee; Ye, Qi; Han, Jie; Meyyappan, M.

    2004-01-01

    There is a strong need for faster, cheaper, and simpler methods for nucleic acid analysis in today s clinical tests. Nanotechnologies can potentially provide solutions to these requirements by integrating nanomaterials with biofunctionalities. Dramatic improvement in the sensitivity and multiplexing can be achieved through the high-degree miniaturization. Here, we present our study in the development of an ultrasensitive label-free electronic chip for DNA/RNA analysis based on carbon nanotube nanoelectrode arrays. A reliable nanoelectrode array based on vertically aligned multi-walled carbon nanotubes (MWNTs) embedded in a SiO2 matrix is fabricated using a bottom-up approach. Characteristic nanoelectrode behavior is observed with a low-density MWNT nanoelectrode array in measuring both the bulk and surface immobilized redox species. The open-end of MWNTs are found to present similar properties as graphite edge-plane electrodes, with a wide potential window, flexible chemical functionalities, and good biocompatibility. A BRCA1 related oligonucleotide probe with 18 bases is covalently functionalized at the open ends of the MWNTs and specifically hybridized with an oligonucleotide target as well as a PCR amplicon. The guanine bases in the target molecules are employed as the signal moieties for the electrochemical measurements. Ru(bpy)3(2+) mediator is used to further amplify the guanine oxidation signal. This technique has been employed for direct electrochemical detection of label-free PCR amplicon through specific hybridization with the BRCAl probe. The detection limit is estimated to be less than approximately 1000 DNA molecules, approaching the limit of the sensitivity by laser-based fluorescence techniques in DNA microarray. This system provides a general electronic platform for rapid molecular diagnostics in applications requiring ultrahigh sensitivity, high-degree of miniaturization, simple sample preparation, and low- cost operation.

  18. Dual-point dual-wavelength fluorescence monitoring of DNA separation in a lab on a chip

    PubMed Central

    Dongre, Chaitanya; van Weerd, Jasper; Bellini, Nicola; Osellame, Roberto; Cerullo, Giulio; van Weeghel, Rob; Hoekstra, Hugo J. W. M.; Pollnau, Markus

    2010-01-01

    We present a simple approach in electrophoretic DNA separation and fluorescent monitoring that allows to identify the insertion or deletion of base-pairs in DNA probe molecules from genetic samples, and to perform intrinsic calibration/referencing for highly accurate DNA analysis. The principle is based on dual-point, dual-wavelength laser-induced fluorescence excitation using one or two excitation windows at the intersection of integrated waveguides and microfluidic channels in an optofluidic chip and a single, color-blind photodetector, resulting in a limit of detection of ~200 pM for single-end-labeled DNA molecules. The approach using a single excitation window is demonstrated experimentally, while the option exploiting two excitation windows is proposed theoretically. PMID:21258504

  19. Application of DNA chip scanning technology for automatic detection of Chlamydia trachomatis and Chlamydia pneumoniae inclusions.

    PubMed

    Bogdanov, Anita; Endrész, Valeria; Urbán, Szabolcs; Lantos, Ildikó; Deák, Judit; Burián, Katalin; Önder, Kamil; Ayaydin, Ferhan; Balázs, Péter; Virok, Dezso P

    2014-01-01

    Chlamydiae are obligate intracellular bacteria that propagate in the inclusion, a specific niche inside the host cell. The standard method for counting chlamydiae is immunofluorescent staining and manual counting of chlamydial inclusions. High- or medium-throughput estimation of the reduction in chlamydial inclusions should be the basis of testing antichlamydial compounds and other drugs that positively or negatively influence chlamydial growth, yet low-throughput manual counting is the common approach. To overcome the time-consuming and subjective manual counting, we developed an automatic inclusion-counting system based on a commercially available DNA chip scanner. Fluorescently labeled inclusions are detected by the scanner, and the image is processed by ChlamyCount, a custom plug-in of the ImageJ software environment. ChlamyCount was able to measure the inclusion counts over a 1-log-unit dynamic range with a high correlation to the theoretical counts. ChlamyCount was capable of accurately determining the MICs of the novel antimicrobial compound PCC00213 and the already known antichlamydial antibiotics moxifloxacin and tetracycline. ChlamyCount was also able to measure the chlamydial growth-altering effect of drugs that influence host-bacterium interaction, such as gamma interferon, DEAE-dextran, and cycloheximide. ChlamyCount is an easily adaptable system for testing antichlamydial antimicrobials and other compounds that influence Chlamydia-host interactions. PMID:24189259

  20. An on-chip thin film photodetector for the quantification of DNA probes and targets in microarrays

    PubMed Central

    Fixe, F.; Chu, V.; Prazeres, D. M. F.; Conde, J. P.

    2004-01-01

    A flat microdevice which incorporates a thin-film amorphous silicon (a-Si:H) photodetector with an upper layer of functionalized SiO2 is used to quantify the density of both immobilized and hybridized DNA oligonucleotides labeled with a fluorophore. The device is based on the photoconductivity of hydrogenated amorphous silicon in a coplanar electrode configuration. Excitation, with near UV/blue light, of a single-stranded DNA molecule tagged with the fluorophore 1-(3-(succinimidyloxycarbonyl)benzyl)-4-(5-(4-methoxyphenyl)oxazol-2-yl) pyridinium bromide (PyMPO), results in the emission of visible light. The emitted light is then converted into an electrical signal in the photodetector, thus allowing the optoelectronic detection of the DNA molecules. The detection limit of the present device is of the order of 1 × 1012 molecules/cm2 and is limited by the efficiency of the filtering of the excitation light. A surface density of 33.5 ± 4.0 pmol/cm2 was measured for DNA covalently immobilized to the functionalized SiO2 thin film and a surface density of 3.7 ± 1.5 pmol/cm2 was measured for the complementary DNA hybridized to the bound DNA. The detection concept explored can enable on-chip electronic data acquisition, improving both the speed and the reliability of DNA microarrays. PMID:15148343

  1. An on-chip thin film photodetector for the quantification of DNA probes and targets in microarrays.

    PubMed

    Fixe, F; Chu, V; Prazeres, D M F; Conde, J P

    2004-01-01

    A flat microdevice which incorporates a thin-film amorphous silicon (a-Si:H) photodetector with an upper layer of functionalized SiO2 is used to quantify the density of both immobilized and hybridized DNA oligonucleotides labeled with a fluorophore. The device is based on the photoconductivity of hydrogenated amorphous silicon in a coplanar electrode configuration. Excitation, with near UV/blue light, of a single-stranded DNA molecule tagged with the fluorophore 1-(3-(succinimidyloxycarbonyl)benzyl)-4-(5-(4-methoxyphenyl)oxazol-2-yl) pyridinium bromide (PyMPO), results in the emission of visible light. The emitted light is then converted into an electrical signal in the photodetector, thus allowing the optoelectronic detection of the DNA molecules. The detection limit of the present device is of the order of 1 x 10(12) molecules/cm2 and is limited by the efficiency of the filtering of the excitation light. A surface density of 33.5 +/- 4.0 pmol/cm2 was measured for DNA covalently immobilized to the functionalized SiO2 thin film and a surface density of 3.7 +/- 1.5 pmol/cm2 was measured for the complementary DNA hybridized to the bound DNA. The detection concept explored can enable on-chip electronic data acquisition, improving both the speed and the reliability of DNA microarrays. PMID:15148343

  2. Chromatin immunoprecipitation (ChIP) coupled to detection by quantitative real-time PCR to study transcription factor binding to DNA in Caenorhabditis elegans

    PubMed Central

    Mukhopadhyay, Arnab; Deplancke, Bart; Walhout, Albertha J M; Tissenbaum, Heidi A

    2009-01-01

    In order to determine how signaling pathways differentially regulate gene expression, it is necessary to identify the interactions between transcription factors (TFs) and their cognate cis-regulatory DNA elements. Here, we have outlined a chromatin immunoprecipitation (ChIP) protocol for use in whole Caenorhabditis elegans extracts. We discuss optimization of the procedure, including growth and harvesting of the worms, formaldehyde fixation, TF immunoprecipitation and analysis of bound sequences through real-time PCR. It takes ∼10–12 d to obtain the worm culture for ChIP; the ChIP procedure is spaced out over a period of 2.5 d with two overnight incubations. PMID:18388953

  3. The Hydrogenase Chip: a tiling oligonucleotide DNA microarray technique for characterizing hydrogen-producing and -consuming microbes in microbial communities

    PubMed Central

    Marshall, Ian PG; Berggren, Dusty RV; Azizian, Mohammad F; Burow, Luke C; Semprini, Lewis; Spormann, Alfred M

    2012-01-01

    We developed a broad-ranging method for identifying key hydrogen-producing and consuming microorganisms through analysis of hydrogenase gene content and expression in complex anaerobic microbial communities. The method is based on a tiling hydrogenase gene oligonucleotide DNA microarray (Hydrogenase Chip), which implements a high number of probes per gene by tiling probe sequences across genes of interest at 1.67 × –2 × coverage. This design favors the avoidance of false positive gene identification in samples of DNA or RNA extracted from complex microbial communities. We applied this technique to interrogate interspecies hydrogen transfer in complex communities in (i) lab-scale reductive dehalogenating microcosms enabling us to delineate key H2-consuming microorganisms, and (ii) hydrogen-generating microbial mats where we found evidence for significant H2 production by cyanobacteria. Independent quantitative PCR analysis on selected hydrogenase genes showed that this Hydrogenase Chip technique is semiquantitative. We also determined that as microbial community complexity increases, specificity must be traded for sensitivity in analyzing data from tiling DNA microarrays. PMID:21993396

  4. “Non-canonical protein-DNA interactions identified by ChIP are not artifacts”: response

    PubMed Central

    2013-01-01

    Background Studies of protein association with DNA on a genome wide scale are possible through methods like ChIP-Chip or ChIP-Seq. Massive problems with false positive signals in our own experiments motivated us to revise the standard ChIP-Chip protocol. Analysis of chromosome wide binding of the alternative sigma factor σ32 in Escherichia coli with this new protocol resulted in detection of only a subset of binding sites found in a previous study by Wade and colleagues. We suggested that the remainder of binding sites detected in the previous study are likely to be false positives. In a recent article the Wade group claimed that our conclusion is wrong and that the disputed sites are genuine σ32 binding sites. They further claimed that the non-detection of these sites in our study was due to low data quality. Results/discussion We respond to the criticism of Wade and colleagues and discuss some general questions of ChIP-based studies. We outline why the quality of our data is sufficient to derive meaningful results. Specific points are: (i) the modifications we introduced into the standard ChIP-Chip protocol do not necessarily result in a low dynamic range, (ii) correlation between ChIP-Chip replicates should not be calculated based on the whole data set as done in transcript analysis, (iii) control experiments are essential for identifying false positives. Suggestions are made how ChIP-based methods could be further optimized and which alternative approaches can be used to strengthen conclusions. Conclusion We appreciate the ongoing discussion about the ChIP-Chip method and hope that it helps other scientist to analyze and interpret their results. The modifications we introduced into the ChIP-Chip protocol are a first step towards reducing false positive signals but there is certainly potential for further optimization. The discussion about the σ32 binding sites in question highlights the need for alternative approaches and further investigation of appropriate

  5. Existing and emerging detection technologies for DNA (Deoxyribonucleic Acid) finger printing, sequencing, bio- and analytical chips: a multidisciplinary development unifying molecular biology, chemical and electronics engineering.

    PubMed

    Kumar Khanna, Vinod

    2007-01-01

    The current status and research trends of detection techniques for DNA-based analysis such as DNA finger printing, sequencing, biochips and allied fields are examined. An overview of main detectors is presented vis-à-vis these DNA operations. The biochip method is explained, the role of micro- and nanoelectronic technologies in biochip realization is highlighted, various optical and electrical detection principles employed in biochips are indicated, and the operational mechanisms of these detection devices are described. Although a diversity of biochips for diagnostic and therapeutic applications has been demonstrated in research laboratories worldwide, only some of these chips have entered the clinical market, and more chips are awaiting commercialization. The necessity of tagging is eliminated in refractive-index change based devices, but the basic flaw of indirect nature of most detection methodologies can only be overcome by generic and/or reagentless DNA sensors such as the conductance-based approach and the DNA-single electron transistor (DNA-SET) structure. Devices of the electrical detection-based category are expected to pave the pathway for the next-generation DNA chips. The review provides a comprehensive coverage of the detection technologies for DNA finger printing, sequencing and related techniques, encompassing a variety of methods from the primitive art to the state-of-the-art scenario as well as promising methods for the future.

  6. Clinical Significance of an HPV DNA Chip Test with Emphasis on HPV-16 and/or HPV-18 Detection in Korean Gynecological Patients

    PubMed Central

    Yeo, Min-Kyung; Lee, Ahwon; Hur, Soo Young; Park, Jong Sup

    2016-01-01

    Background: Human papillomavirus (HPV) is a major risk factor for cervical cancer. Methods: We evaluated the clinical significance of the HPV DNA chip genotyping assay (MyHPV chip, Mygene Co.) compared with the Hybrid Capture 2 (HC2) chemiluminescent nucleic acid hybridization kit (Digene Corp.) in 867 patients. Results: The concordance rate between the MyHPV chip and HC2 was 79.4% (kappa coefficient, κ = 0.55). The sensitivity and specificity of both HPV tests were very similar (approximately 85% and 50%, respectively). The addition of HPV result (either MyHPV chip or HC2) to cytology improved the sensitivity (95%, each) but reduced the specificity (approximately 30%, each) compared with the HPV test or cytology alone. Based on the MyHPV chip results, the odds ratio (OR) for ≥ high-grade squamous intraepithelial lesions (HSILs) was 9.9 in the HPV-16/18 (+) group and 3.7 in the non-16/18 high-risk (HR)-HPV (+) group. Based on the HC2 results, the OR for ≥ HSILs was 5.9 in the HR-HPV (+) group. When considering only patients with cytological diagnoses of “negative for intraepithelial lesion or malignancy” and “atypical squamous cell or atypical glandular cell,” based on the MyHPV chip results, the ORs for ≥ HSILs were 6.8 and 11.7, respectively, in the HPV-16/18 (+) group. Conclusions: The sensitivity and specificity of the MyHPV chip test are similar to the HC2. Detecting HPV-16/18 with an HPV DNA chip test, which is commonly used in many Asian countries, is useful in assessing the risk of high-grade cervical lesions. PMID:27345180

  7. Detection and Genotyping of Human Papillomavirus DNA in Formalin-Fixed Paraffin-Embedded Specimens with the HPV Direct Flow CHIP System

    PubMed Central

    Herraez-Hernandez, Elsa; Preda, Ovidiu; Alonso, Sonia; Pardo, Rosario Serrano; Olmo, Asuncion

    2013-01-01

    The novel HPV Direct Flow CHIP commercial system for Human Papillomavirus (HPV) genotyping is based on rapid PCR and automatic reverse dot blot hybridization to genotype-specific probes, allowing the detection of 36 HPV genotypes. This study examined the performance of HPV Direct Flow CHIP in formalin-fixed paraffin-embedded (FFPE) samples (n= 99). Each sample was analyzed both by Direct PCR, using crude cell extracts without DNA purification, and by conventional PCR, using purified DNA. Pair-wise analysis of the results demonstrated strong concordance between the results obtained with the two protocols, although a slightly higher rate of multiple infections was detected by conventional PCR. In summary, HPV Direct Flow CHIP achieves effective HPV detection from FFPE samples with both Direct PCR and Conventional PCR protocols. PMID:24222806

  8. Improved sandwich-hybridization assay for an electrical DNA-chip-based monitoring of bioprocess-relevant marker genes.

    PubMed

    Pioch, Daniel; Jürgen, Britta; Evers, Stefan; Maurer, Karl-Heinz; Hecker, Michael; Schweder, Thomas

    2008-03-01

    Recently, it was shown that electrical DNA-chips in connection with a magnetic bead-based sandwich-hybridization assay can be a suitable alternative for the analysis of gene expression by monitoring the respective mRNA levels. In this study, we established an improved assay which allowed for a significantly shortened but sensitive detection of specific mRNAs. These improvements include the change to a solution-based sandwich-hybridization and the rearrangement of the used oligonucleotide probes. The introduction of a second labeled detection probe further increased the hybridization signals and, in turn, leads to a substantial time reduction of the detection protocol. The presented solution-based sandwich-hybridization protocol for the electrochemical detection of specific mRNAs requires about 60 min and the whole procedure, including sampling, cell disruption, and RNA isolation, approx. 80 min. The assay of this study was verified by nutrient-limited growth experiments and the analysis of selected starvation marker genes of the industrial host Bacillus licheniformis. The expression profiles determined with the electrical chip and the optimized protocol were, in most cases, comparable with the profiles determined by real-time RT-PCR measurements.

  9. Genome-Wide Profiling of Yeast DNA:RNA Hybrid Prone Sites with DRIP-Chip

    PubMed Central

    Lu, Phoebe Y. T.; Luo, Zongli; Hamza, Akil; Kobor, Michael S.; Stirling, Peter C.; Hieter, Philip

    2014-01-01

    DNA:RNA hybrid formation is emerging as a significant cause of genome instability in biological systems ranging from bacteria to mammals. Here we describe the genome-wide distribution of DNA:RNA hybrid prone loci in Saccharomyces cerevisiae by DNA:RNA immunoprecipitation (DRIP) followed by hybridization on tiling microarray. These profiles show that DNA:RNA hybrids preferentially accumulated at rDNA, Ty1 and Ty2 transposons, telomeric repeat regions and a subset of open reading frames (ORFs). The latter are generally highly transcribed and have high GC content. Interestingly, significant DNA:RNA hybrid enrichment was also detected at genes associated with antisense transcripts. The expression of antisense-associated genes was also significantly altered upon overexpression of RNase H, which degrades the RNA in hybrids. Finally, we uncover mutant-specific differences in the DRIP profiles of a Sen1 helicase mutant, RNase H deletion mutant and Hpr1 THO complex mutant compared to wild type, suggesting different roles for these proteins in DNA:RNA hybrid biology. Our profiles of DNA:RNA hybrid prone loci provide a resource for understanding the properties of hybrid-forming regions in vivo, extend our knowledge of hybrid-mitigating enzymes, and contribute to models of antisense-mediated gene regulation. A summary of this paper was presented at the 26th International Conference on Yeast Genetics and Molecular Biology, August 2013. PMID:24743342

  10. Genome-wide profiling of yeast DNA:RNA hybrid prone sites with DRIP-chip.

    PubMed

    Chan, Yujia A; Aristizabal, Maria J; Lu, Phoebe Y T; Luo, Zongli; Hamza, Akil; Kobor, Michael S; Stirling, Peter C; Hieter, Philip

    2014-04-01

    DNA:RNA hybrid formation is emerging as a significant cause of genome instability in biological systems ranging from bacteria to mammals. Here we describe the genome-wide distribution of DNA:RNA hybrid prone loci in Saccharomyces cerevisiae by DNA:RNA immunoprecipitation (DRIP) followed by hybridization on tiling microarray. These profiles show that DNA:RNA hybrids preferentially accumulated at rDNA, Ty1 and Ty2 transposons, telomeric repeat regions and a subset of open reading frames (ORFs). The latter are generally highly transcribed and have high GC content. Interestingly, significant DNA:RNA hybrid enrichment was also detected at genes associated with antisense transcripts. The expression of antisense-associated genes was also significantly altered upon overexpression of RNase H, which degrades the RNA in hybrids. Finally, we uncover mutant-specific differences in the DRIP profiles of a Sen1 helicase mutant, RNase H deletion mutant and Hpr1 THO complex mutant compared to wild type, suggesting different roles for these proteins in DNA:RNA hybrid biology. Our profiles of DNA:RNA hybrid prone loci provide a resource for understanding the properties of hybrid-forming regions in vivo, extend our knowledge of hybrid-mitigating enzymes, and contribute to models of antisense-mediated gene regulation. A summary of this paper was presented at the 26th International Conference on Yeast Genetics and Molecular Biology, August 2013. PMID:24743342

  11. Simultaneous detection of duplex DNA oligonucleotides using a SERS-based micro-network gradient chip.

    PubMed

    Choi, Namhyun; Lee, Kangsun; Lim, Dong Woo; Lee, Eun Kyu; Chang, Soo-Ik; Oh, Kwang W; Choo, Jaebum

    2012-12-21

    We report the development of a programmable surface-enhanced Raman scattering (SERS)-based micro-network gradient platform to simultaneously detect two different types of DNA oligomer mixtures. The utility of this platform was demonstrated by quantitative analysis of two breast cancer-related (BRCA1) DNA oligomer mixtures. To generate on-demand concentration gradients, the microfluidic circuit was designed using an electric-hydraulic analogy. Then a multi-gradient microfluidic channel was fabricated based on the theoretical design of the concentration control module. These micro-network structures automatically produce a series of different concentration gradients by continuously mixing Cy3-labeled DNA oligomers (BRAC1-Mutation) with TAMRA-labeled DNA oligomer (BRAC1-Wild). The SERS signals for different ratios of duplex DNA oligomer mixtures, adsorbed on the surface of silver nanoparticles, were measured under flowing conditions. Total analysis time from serial mixing to SERS detection takes less than 10 min because all experimental conditions are automatically controlled inside the exquisitely designed microfluidic channel. This novel SERS-based DNA sensing technology in a micro-network gradient channel is expected to be a powerful analytical tool to simultaneously detect multiple DNA oligomer mixtures.

  12. Chip, Chip, Hooray!

    ERIC Educational Resources Information Center

    Kelly, Susan

    2001-01-01

    Presents a science laboratory using different brands of potato chips in which students test their oiliness, size, thickness, saltiness, quality, and cost, then analyze the results to determine the best chip. Gives a brief history of potato chips. (YDS)

  13. First all-in-one diagnostic tool for DNA intelligence: genome-wide inference of biogeographic ancestry, appearance, relatedness, and sex with the Identitas v1 Forensic Chip.

    PubMed

    Keating, Brendan; Bansal, Aruna T; Walsh, Susan; Millman, Jonathan; Newman, Jonathan; Kidd, Kenneth; Budowle, Bruce; Eisenberg, Arthur; Donfack, Joseph; Gasparini, Paolo; Budimlija, Zoran; Henders, Anjali K; Chandrupatla, Hareesh; Duffy, David L; Gordon, Scott D; Hysi, Pirro; Liu, Fan; Medland, Sarah E; Rubin, Laurence; Martin, Nicholas G; Spector, Timothy D; Kayser, Manfred

    2013-05-01

    When a forensic DNA sample cannot be associated directly with a previously genotyped reference sample by standard short tandem repeat profiling, the investigation required for identifying perpetrators, victims, or missing persons can be both costly and time consuming. Here, we describe the outcome of a collaborative study using the Identitas Version 1 (v1) Forensic Chip, the first commercially available all-in-one tool dedicated to the concept of developing intelligence leads based on DNA. The chip allows parallel interrogation of 201,173 genome-wide autosomal, X-chromosomal, Y-chromosomal, and mitochondrial single nucleotide polymorphisms for inference of biogeographic ancestry, appearance, relatedness, and sex. The first assessment of the chip's performance was carried out on 3,196 blinded DNA samples of varying quantities and qualities, covering a wide range of biogeographic origin and eye/hair coloration as well as variation in relatedness and sex. Overall, 95 % of the samples (N = 3,034) passed quality checks with an overall genotype call rate >90 % on variable numbers of available recorded trait information. Predictions of sex, direct match, and first to third degree relatedness were highly accurate. Chip-based predictions of biparental continental ancestry were on average ~94 % correct (further support provided by separately inferred patrilineal and matrilineal ancestry). Predictions of eye color were 85 % correct for brown and 70 % correct for blue eyes, and predictions of hair color were 72 % for brown, 63 % for blond, 58 % for black, and 48 % for red hair. From the 5 % of samples (N = 162) with <90 % call rate, 56 % yielded correct continental ancestry predictions while 7 % yielded sufficient genotypes to allow hair and eye color prediction. Our results demonstrate that the Identitas v1 Forensic Chip holds great promise for a wide range of applications including criminal investigations, missing person investigations, and for national security

  14. LoMA-B: a simple and versatile lab-on-a-chip system based on single-channel bisulfite conversion for DNA methylation analysis.

    PubMed

    Yoon, Jaeyun; Park, Mi Kyoung; Lee, Tae Yoon; Yoon, Yong-Jin; Shin, Yong

    2015-09-01

    Miniaturized lab-on-a-chip (LOC) systems have been developed for genetic and epigenetic analyses in clinical applications because of advantages such as reduced sample size and reagent consumption, rapid processing speed, simplicity, and enhanced sensitivity. Despite tremendous efforts made towards developing LOC systems for use in the clinical setting, the development of LOC systems to analyze DNA methylation, which is an emerging epigenetic marker causing the abnormal silencing of genes including tumor suppressor genes, is still challenging because of the gold standard methods involving a bisulfite conversion step. Existing bisulfite conversion-based techniques are not suitable for clinical use due to their long processing time, labor intensiveness, and the purification steps involved. Here, we present a lab-on-a-chip system for DNA methylation analysis based on bisulfite conversion (LoMA-B), which couples a sample pre-processing module for on-chip bisulfite conversion and a label-free, real-time detection module for rapid analysis of DNA methylation status using an isothermal DNA amplification/detection technique. The methylation status of the RARβ gene in human genomic DNA extracted from MCF-7 cells was analyzed by the LoMA-B system within 80 min (except 16 h for sensor preparation) compared to conventional MS-PCR within 24 h. Furthermore, the LoMA-B system is highly sensitive and can detect as little as 1% methylated DNA in a methylated/unmethylated cell mixture. Therefore, the LoMA-B system is an efficient diagnostic tool for the simple, versatile, and quantitative evaluation of DNA methylation patterns for clinical applications.

  15. Microfluidics-Based Lab-on-Chip Systems in DNA-Based Biosensing: An Overview

    PubMed Central

    Dutse, Sabo Wada; Yusof, Nor Azah

    2011-01-01

    Microfluidics-based lab-on-chip (LOC) systems are an active research area that is revolutionising high-throughput sequencing for the fast, sensitive and accurate detection of a variety of pathogens. LOCs also serve as portable diagnostic tools. The devices provide optimum control of nanolitre volumes of fluids and integrate various bioassay operations that allow the devices to rapidly sense pathogenic threat agents for environmental monitoring. LOC systems, such as microfluidic biochips, offer advantages compared to conventional identification procedures that are tedious, expensive and time consuming. This paper aims to provide a broad overview of the need for devices that are easy to operate, sensitive, fast, portable and sufficiently reliable to be used as complementary tools for the control of pathogenic agents that damage the environment. PMID:22163925

  16. Microfluidics-based lab-on-chip systems in DNA-based biosensing: an overview.

    PubMed

    Dutse, Sabo Wada; Yusof, Nor Azah

    2011-01-01

    Microfluidics-based lab-on-chip (LOC) systems are an active research area that is revolutionising high-throughput sequencing for the fast, sensitive and accurate detection of a variety of pathogens. LOCs also serve as portable diagnostic tools. The devices provide optimum control of nanolitre volumes of fluids and integrate various bioassay operations that allow the devices to rapidly sense pathogenic threat agents for environmental monitoring. LOC systems, such as microfluidic biochips, offer advantages compared to conventional identification procedures that are tedious, expensive and time consuming. This paper aims to provide a broad overview of the need for devices that are easy to operate, sensitive, fast, portable and sufficiently reliable to be used as complementary tools for the control of pathogenic agents that damage the environment.

  17. Liquid sensor based bio-chip for DNA analysis of cancer using photonic crystal

    NASA Astrophysics Data System (ADS)

    Patil, Harshada; Nischitha, R.; Indumathi, T. S.; Sharan, Preeta

    2015-07-01

    Silicon photonics is poised to revolutionize bio-sensing applications, specifically in medical diagnostics. The need for cost effective and reliable bio-sensors in medical applications is an ever growing and everlasting one. In this synopsis we have designed a 2-D hexagonal photonic crystal ring resonator based bio-sensor that is able to detect lung cancer from blood. Simulation and analysis has been done for normal DNA and the cancer affected DNA in blood. The intensity level of transmission spectrum has been observed. Finite Difference Time Domain (FDTD) method is used for analysis. MEEP (MIT Electromagnetic Equation Propagation) tool and RSOFT Photonic Suite CAD tool are used designing the photonic crystal sensor. The results show that for small changes in the refractive index of the input samples there is a significant shift in wavelength and amplitude. Thus the sensor is highly sensitive for change in refractive index and hence differentiating normal and cancer affected DNA.

  18. Ternary DNA chip based on a novel thymine spacer group chemistry.

    PubMed

    Yang, Yanli; Yildiz, Umit Hakan; Peh, Jaime; Liedberg, Bo

    2015-01-01

    A novel thymine-based surface chemistry suitable for label-free electrochemical DNA detection is described. It involves a simple two-step sequential process: immobilization of 9-mer thymine-terminated probe DNAs followed by backfilling with 9-mer thymine-based spacers (T9). As compared to commonly used organic spacer groups like 2-mercaptoethanol, 3-mercapto-1-propanol and 6-mercapto-1-hexanol, the 9-mer thymine-based spacers offer a 10-fold improvement in discriminating between complementary and non-complementary target hybridization, which is due mainly to facilitated transport of the redox probes through the probe-DNA/T9 layers. Electrochemical measurements, complemented with Surface Plasmon Resonance (SPR) and Quartz Crystal Microbalance (QCM-D) binding analyses, reveal that optimum selectivity between complementary and non-complementary hybridization is obtained for a sensing surface prepared using probe-DNA and backfiller T9 at equimolar concentration (1:1). At this particular ratio, the probe-DNAs are preferentially oriented and easily accessible to yield a sensing surface with favorable hybridization and electron transfer characteristics. Our findings suggest that oligonucleotide-based spacer groups offer an attractive alternative to short organic thiol spacers in the design of future DNA biochips. PMID:25465760

  19. Carbon Nanotube Nanoelectrode Array as an Electronic Chip for Ultrasensitive Label-free DNA Detection

    NASA Technical Reports Server (NTRS)

    Li, Jun; Koehne, Jessica; Chen, Hua; Cassell, Alan; Ng, Hou Tee; Fan, Wendy; Ye, Qi; Han, Jie; Meyyappan, M.

    2003-01-01

    A reliable nanoelectrode array based on vertically aligned multi-walled carbon nanotubes (MWNTs) embedded in SiO2 is used for ultrasensitive DNA detection. Characteristic nanoelectrode behavior is observed using low-density MWNT arrays for measuring both bulk and surface immobilized redox species such as K4Fe(CN)6 and ferrocene derivatives. The open-end of MWNTs are found to present similar properties as graphite edge-plane electrodes with wide potential window, flexible chemical functionalities, and good biocompatibility. BRCA1 related oligonucleotide probes with 18 bp are selectively functionalized at the open ends of the nanotube array and specifically hybridized with oligonucleotide targets incorporated with a polyG tag. The guanine groups are employed as the signal moieties in the electrochemical measurements. R(bpy)(sup 2+, sub 3) mediator is used to further amplify the guanine oxidation signal. The hybridization of sub-attomoles of DNA targets is detected electrochemically by combining the MWNT nanoelectrode array with the R(bpy)(sup 2+, sub 3) amplification mechanism. This technique was employed for direct electrochemical detection of label-free PCR amplicon from a healthy donor through specific hybridization with the BRCA1 probe. The detection limit is estimated to be less than 1000 DNA molecules since abundant guanine bases in the PCR amplicon provides a large signal. This system provides a general platform for rapid molecular diagnostics in applications requiring ultrahigh sensitivity, high-degree of miniaturization, and simple sample preparation, and low-cost operation.

  20. Dynamic chromatin remodelling of ciliate macronuclear DNA as determined by an optimized chromatin immunoprecipitation (ChIP) method for Paramecium tetraurelia.

    PubMed

    Cheaib, Miriam; Simon, Martin

    2013-03-01

    We report the detailed evaluation of crucial parameters for chromatin immunoprecipitation (ChIP) of macronuclear DNA in the unicellular eukaryote Paramecium tetraurelia. Optimized parameters include crosslinking conditions, chromatin sonication and antibody titration thus providing a detailed protocol for successful ChIP in P. tetraurelia. As this ciliate is bacterivorous and RNAi by feeding represents a powerful tool for analysis of gene function, we moreover determined the effects of ingested nucleic acids by food bacteria. Feasibility of our protocol is demonstrated by characterisation of chromatin remodelling at promoters of cytosolic HSP70 isoforms during transcriptional activation under heat shock conditions by analyzing RNA abundance, nucleosome occupancy and levels of H3 lysine 9 acetylation.

  1. Protein analysis by time-resolved measurements with an electro-switchable DNA chip

    PubMed Central

    Langer, Andreas; Hampel, Paul A.; Kaiser, Wolfgang; Knezevic, Jelena; Welte, Thomas; Villa, Valentina; Maruyama, Makiko; Svejda, Matej; Jähner, Simone; Fischer, Frank; Strasser, Ralf; Rant, Ulrich

    2013-01-01

    Measurements in stationary or mobile phases are fundamental principles in protein analysis. Although the immobilization of molecules on solid supports allows for the parallel analysis of interactions, properties like size or shape are usually inferred from the molecular mobility under the influence of external forces. However, as these principles are mutually exclusive, a comprehensive characterization of proteins usually involves a multi-step workflow. Here we show how these measurement modalities can be reconciled by tethering proteins to a surface via dynamically actuated nanolevers. Short DNA strands, which are switched by alternating electric fields, are employed as capture probes to bind target proteins. By swaying the proteins over nanometre amplitudes and comparing their motional dynamics to a theoretical model, the protein diameter can be quantified with Angström accuracy. Alterations in the tertiary protein structure (folding) and conformational changes are readily detected, and even post-translational modifications are revealed by time-resolved molecular dynamics measurements. PMID:23839273

  2. Hepatology in the 21st century. Gene transfer, hepatocyte transplantation, DNA chips, cyberspace and ... a friendly hospital.

    PubMed

    Jansen, P L

    1999-12-01

    What to expect for hepatology in the 21st century? If science is allowed to proceed at its current rate, expectations can hardly be underestimated. Bound by the present day's limitations we are only able to see a glimpse of what could be available 100 years from now. For the next few decades, the global eradication of viral hepatitis will be on the agenda. For the treatment of inherited and acquired metabolic, toxic and immune liver disease, targeted drugs, genes and antisense oligonucleotides will be added to our therapeutic repertoire. The completion of the human genome project in 2003 will have far-reaching consequences: the widespread use of prenatal diagnosis, using DNA chip technology, may be expected to cause a dramatic decrease in the incidence of inherited diseases. Liver cirrhosis, hepatocellular carcinoma and inborn errors of metabolism may be treated by gene transfer or gene repair therapy. Although eventually these developments may decrease the need for organ transplantation, this by no means is the case yet and no solution is available for an increased demand and a decreased supply of organs. In the long run, diseases caused by multi-drug-resistant infectious agents and diseases associated with the abuse of alcohol and drugs are expected to become major problems. The future of university-based research is uncertain. The staggering costs of research and limited career possibilities may force universities to the limited task of higher education, with as a result biotech companies, shareholders and corporate finance ruling the scientific waves in the next century. The 21st century patient will know the way in cyberspace and will go shopping for the best doctor, for the best treatment and for the best, or friendliest, hospital. PMID:10628176

  3. MTB-DR-RIF 9G test: Detection and discrimination of tuberculosis and multi-drug resistant tuberculosis strains.

    PubMed

    Song, Keum-Soo; Nimse, Satish Balasaheb; Cho, Nam Hoon; Sung, Nackmoon; Kim, Hee-Jin; Yang, Jeongseong; Kim, Taisun

    2015-12-01

    This report describes the evaluation of the novel MTB-DR-RIF 9G test for the accurate detection and discrimination of Mycobacterium tuberculosis (MTB) and rifampicin-resistant M. tuberculosis (MTB-DR-RIF) in the clinical samples. The procedure included the amplification of a nucleotide fragment of the rpoB gene of the MTB and MTB-DR-RIF strains and their hybridization with the immobilized probes. The MTB-DR-RIF 9G test was evaluated for its ability to detect and discriminate MTB and MTB-DR-RIF strains in 113 known clinical samples. The accuracy of the MTB-DR-RIF 9G test was determined by comparing its results with sequencing analysis and drug susceptibility testing. The sensitivity and specificity of the MTB-DR-RIF 9G test at 95% confidence interval were found to be 95.4% (89.5-98.5) and 100% (69.2-100), respectively. The positive predictive value and negative predictive value of the MTB-DR-RIF 9G test at 95% confidence interval were found to be 100% (85.0-95.9) and 66.7% (38.4-88.18), respectively. Sequencing analysis of all samples indicated that the mutations present in the regions identified with the MTB-DR-RIF 9G assay can be detected accurately.

  4. On-chip FRET Graphene Oxide Aptasensor: Quantitative Evaluation of Enhanced Sensitivity by Aptamer with a Double-stranded DNA Spacer.

    PubMed

    Ueno, Yuko; Furukawa, Kazuaki; Tin, Andrew; Hibino, Hiroki

    2015-01-01

    We propose a molecular design for a biomolecular probe to realize an on-chip graphene oxide (GO) aptasensor with enhanced sensitivity. Here, GO works as an excellent acceptor for fluorescence resonance energy transfer. We inserted a rigid double-stranded DNA as a spacer between the GO surface and the aptamer sequence to extend the distance between a fluorescence dye and the GO surface during molecular recognition. We examined the dependence of the sensitivity on the length of the spacer quantitatively by using a 2×2 linear-array aptasensor. We used the modified aptamer with 10 and 30 base pair (bp) double-stranded DNA spacers. The signal with a 30bp-spacer was about twice as strong that with a 10bp-spacer as regards both thrombin and prostate specific antigen detections. The improvement in the sensitivity was supported by a model calculation that estimated the effect of spacer length on fluorescence recovery efficiency. PMID:26353952

  5. Sensitive and accurate identification of protein–DNA binding events in ChIP-chip assays using higher order derivative analysis

    PubMed Central

    Barrett, Christian L.; Cho, Byung-Kwan

    2011-01-01

    Immuno-precipitation of protein–DNA complexes followed by microarray hybridization is a powerful and cost-effective technology for discovering protein–DNA binding events at the genome scale. It is still an unresolved challenge to comprehensively, accurately and sensitively extract binding event information from the produced data. We have developed a novel strategy composed of an information-preserving signal-smoothing procedure, higher order derivative analysis and application of the principle of maximum entropy to address this challenge. Importantly, our method does not require any input parameters to be specified by the user. Using genome-scale binding data of two Escherichia coli global transcription regulators for which a relatively large number of experimentally supported sites are known, we show that ∼90% of known sites were resolved to within four probes, or ∼88 bp. Over half of the sites were resolved to within two probes, or ∼38 bp. Furthermore, we demonstrate that our strategy delivers significant quantitative and qualitative performance gains over available methods. Such accurate and sensitive binding site resolution has important consequences for accurately reconstructing transcriptional regulatory networks, for motif discovery, for furthering our understanding of local and non-local factors in protein–DNA interactions and for extending the usefulness horizon of the ChIP-chip platform. PMID:21051353

  6. Combining combing and secondary ion mass spectrometry to study DNA on chips using (13)C and (15)N labeling.

    PubMed

    Cabin-Flaman, Armelle; Monnier, Anne-Francoise; Coffinier, Yannick; Audinot, Jean-Nicolas; Gibouin, David; Wirtz, Tom; Boukherroub, Rabah; Migeon, Henri-Noël; Bensimon, Aaron; Jannière, Laurent; Ripoll, Camille; Norris, Victor

    2016-01-01

    Dynamic secondary ion mass spectrometry ( D-SIMS) imaging of combed DNA - the combing, imaging by SIMS or CIS method - has been developed previously using a standard NanoSIMS 50 to reveal, on the 50 nm scale, individual DNA fibers labeled with different, non-radioactive isotopes in vivo and to quantify these isotopes. This makes CIS especially suitable for determining the times, places and rates of DNA synthesis as well as the detection of the fine-scale re-arrangements of DNA and of molecules associated with combed DNA fibers. Here, we show how CIS may be extended to (13)C-labeling via the detection and quantification of the (13)C (14)N (-) recombinant ion and the use of the (13)C: (12)C ratio, we discuss how CIS might permit three successive labels, and we suggest ideas that might be explored using CIS. PMID:27429742

  7. Combining combing and secondary ion mass spectrometry to study DNA on chips using 13C and 15N labeling

    PubMed Central

    Cabin-Flaman, Armelle; Monnier, Anne-Francoise; Coffinier, Yannick; Audinot, Jean-Nicolas; Gibouin, David; Wirtz, Tom; Boukherroub, Rabah; Migeon, Henri-Noël; Bensimon, Aaron; Jannière, Laurent; Ripoll, Camille; Norris, Victor

    2016-01-01

    Dynamic secondary ion mass spectrometry ( D-SIMS) imaging of combed DNA – the combing, imaging by SIMS or CIS method – has been developed previously using a standard NanoSIMS 50 to reveal, on the 50 nm scale, individual DNA fibers labeled with different, non-radioactive isotopes in vivo and to quantify these isotopes. This makes CIS especially suitable for determining the times, places and rates of DNA synthesis as well as the detection of the fine-scale re-arrangements of DNA and of molecules associated with combed DNA fibers. Here, we show how CIS may be extended to 13C-labeling via the detection and quantification of the 13C 14N - recombinant ion and the use of the 13C: 12C ratio, we discuss how CIS might permit three successive labels, and we suggest ideas that might be explored using CIS. PMID:27429742

  8. Genome-wide DNA methylation detection by MethylCap-seq and Infinium HumanMethylation450 BeadChips: an independent large-scale comparison

    PubMed Central

    De Meyer, Tim; Bady, Pierre; Trooskens, Geert; Kurscheid, Sebastian; Bloch, Jocelyne; Kros, Johan M.; Hainfellner, Johannes A.; Stupp, Roger; Delorenzi, Mauro; Hegi, Monika E.; Van Criekinge, Wim

    2015-01-01

    Two cost-efficient genome-scale methodologies to assess DNA-methylation are MethylCap-seq and Illumina’s Infinium HumanMethylation450 BeadChips (HM450). Objective information regarding the best-suited methodology for a specific research question is scant. Therefore, we performed a large-scale evaluation on a set of 70 brain tissue samples, i.e. 65 glioblastoma and 5 non-tumoral tissues. As MethylCap-seq coverages were limited, we focused on the inherent capacity of the methodology to detect methylated loci rather than a quantitative analysis. MethylCap-seq and HM450 data were dichotomized and performances were compared using a gold standard free Bayesian modelling procedure. While conditional specificity was adequate for both approaches, conditional sensitivity was systematically higher for HM450. In addition, genome-wide characteristics were compared, revealing that HM450 probes identified substantially fewer regions compared to MethylCap-seq. Although results indicated that the latter method can detect more potentially relevant DNA-methylation, this did not translate into the discovery of more differentially methylated loci between tumours and controls compared to HM450. Our results therefore indicate that both methodologies are complementary, with a higher sensitivity for HM450 and a far larger genome-wide coverage for MethylCap-seq, but also that a more comprehensive character does not automatically imply more significant results in biomarker studies. PMID:26482909

  9. SR proteins SRp20 and 9G8 contribute to efficient export of herpes simplex virus 1 mRNAs.

    PubMed

    Escudero-Paunetto, Laurimar; Li, Ling; Hernandez, Felicia P; Sandri-Goldin, Rozanne M

    2010-06-01

    Herpes simplex virus 1 (HSV-1) mRNAs are exported to the cytoplasm through the export receptor TAP/NFX1. HSV-1 multifunctional protein ICP27 interacts with TAP/NXF1, binds viral RNAs, and is required for efficient viral RNA export. In ICP27 mutant infections, viral RNA export is reduced but not ablated, indicating that other export adaptors can aid in viral RNA export. Export adaptor protein Aly/REF is recruited to viral replication compartments, however, Aly/REF knockdown has little effect on viral RNA export. SR proteins SRp20 and 9G8 interact with TAP/NXF1 and mediate export of some cellular RNAs. We report that siRNA knockdown of SRp20 or 9G8 resulted in about a 10 fold decrease in virus yields and in nuclear accumulation of polyA+ RNA. In infected cells depleted of SRp20, newly transcribed Bromouridine-labeled RNA also accumulated in the nucleus. We conclude that SRp20 and 9G8 contribute to HSV-1 RNA export. PMID:20227104

  10. SR proteins SRp20 and 9G8 contribute to efficient export of herpes simplex virus 1 mRNAs

    SciTech Connect

    Escudero-Paunetto, Laurimar; Li Ling; Hernandez, Felicia P.; Sandri-Goldin, Rozanne M.

    2010-06-05

    Herpes simplex virus 1 (HSV-1) mRNAs are exported to the cytoplasm through the export receptor TAP/NFX1. HSV-1 multifunctional protein ICP27 interacts with TAP/NXF1, binds viral RNAs, and is required for efficient viral RNA export. In ICP27 mutant infections, viral RNA export is reduced but not ablated, indicating that other export adaptors can aid in viral RNA export. Export adaptor protein Aly/REF is recruited to viral replication compartments, however, Aly/REF knockdown has little effect on viral RNA export. SR proteins SRp20 and 9G8 interact with TAP/NXF1 and mediate export of some cellular RNAs. We report that siRNA knockdown of SRp20 or 9G8 resulted in about a 10 fold decrease in virus yields and in nuclear accumulation of poly(A+) RNA. In infected cells depleted of SRp20, newly transcribed Bromouridine-labeled RNA also accumulated in the nucleus. We conclude that SRp20 and 9G8 contribute to HSV-1 RNA export.

  11. Screening of DNA aptamers against myoglobin using a positive and negative selection units integrated microfluidic chip and its biosensing application.

    PubMed

    Wang, Qing; Liu, Wei; Xing, Yuqian; Yang, Xiaohai; Wang, Kemin; Jiang, Rui; Wang, Pei; Zhao, Qing

    2014-07-01

    An aptamer screening method using a positive and negative selection units integrated microfluidic chip was introduced. Here, myoglobin (Myo), one of the early markers to increase after acute myocardial infarction, was used as the model. After 7-round selection, the aptamers, which exhibited dissociation constants (K(d)) in the nanomolar range (from 4.93 to 6.38 nM), were successfully obtained using a positive and negative selection units integrated microfluidic chip. The aptamer with the highest affinity (K(d) = 4.93 nM) was then used for the fabrication of a label-free supersandwich electrochemical biosensor for Myo detection based on target-induced aptamer displacement. The detection limit of this aptamer-based electrochemical biosensor was 10 pM, which was significantly lower than that of those previous antibody-based biosensors for Myo detection. This work may not only develop a strategy for screening aptamer but also offer promising alternatives to the traditional analytical and immunological methods for Myo detection.

  12. Gene Assembly from Chip-Synthesized Oligonucleotides

    PubMed Central

    Eroshenko, Nikolai; Kosuri, Sriram; Marblestone, Adam H; Conway, Nicholas; Church, George M.

    2012-01-01

    De novo synthesis of long double-stranded DNA constructs has a myriad of applications in biology and biological engineering. However, its widespread adoption has been hindered by high costs. Cost can be significantly reduced by using oligonucleotides synthesized on high-density DNA chips. However, most methods for using off-chip DNA for gene synthesis have failed to scale due to the high error rates, low yields, and high chemical complexity of the chip-synthesized oligonucleotides. We have recently demonstrated that some commercial DNA chip manufacturers have improved error rates, and that the issues of chemical complexity and low yields can be solved by using barcoded primers to accurately and efficiently amplify subpools of oligonucleotides. This article includes protocols for computationally designing the DNA chip, amplifying the oligonucleotide subpools, and assembling 500-800 basepair (bp) constructs. PMID:25077042

  13. Analysis of sequence variations in several human genes using phosphoramidite bond DNA fragmentation and chip-based MALDI-TOF.

    PubMed

    Smylie, Kevin J; Cantor, Charles R; Denissenko, Mikhail F

    2004-01-01

    The challenge in the postgenome era is to measure sequence variations over large genomic regions in numerous patient samples. This massive amount of work can only be completed if more accurate, cost-effective, and high-throughput solutions become available. Here we describe a novel DNA fragmentation approach for single nucleotide polymorphism (SNP) discovery and sequence validation. The base-specific cleavage is achieved by creating primer extension products, in which acid-labile phosphoramidite (P-N) bonds replace the 5' phosphodiester bonds of newly incorporated pyrimidine nucleotides. Sequence variations are detected by hydrolysis of this acid-labile bond and MALDI-TOF analysis of the resulting fragments. In this study, we developed a robust protocol for P-N-bond fragmentation and investigated additional ways to improve its sensitivity and reproducibility. We also present the analysis of several human genomic targets ranging from 100-450 bp in length. By using a semiautomated sample processing protocol, we investigated an array of SNPs within a 240-bp segment of the NFKBIA gene in 48 human DNA samples. We identified and measured frequencies for the two common SNPs in the 3'UTR of NFKBIA (separated by 123 bp) and then confirmed these values in an independent genotyping experiment. The calculated allele frequencies in white and African American groups differed significantly, yet both fit Hardy-Weinberg expectations. This demonstrates the utility and effectiveness of PN-bond DNA fragmentation and subsequent MALDI-TOF MS analysis for the high-throughput discovery and measurement of sequence variations in fragments up to 0.5 kb in length in multiple human blood DNA samples.

  14. Interspecies hybridization on DNA resequencing microarrays: efficiency of sequence recovery and accuracy of SNP detection in human, ape, and codfish mitochondrial DNA genomes sequenced on a human-specific MitoChip

    PubMed Central

    Flynn, Sarah MC; Carr, Steven M

    2007-01-01

    data suggest that interspecific cross-hybridization will not interfere with the accurate recovery of species-specific data from multispecies microarrays, provided that the species' DNA sequences differ by > 20% (mean of 5b differences per 25b oligo). Recovery of DNA sequence data from multiple, distantly-related species on a single multiplex gene chip should be a practical, highly-parallel method for investigating genomic biodiversity. PMID:17894875

  15. DNA.

    ERIC Educational Resources Information Center

    Felsenfeld, Gary

    1985-01-01

    Structural form, bonding scheme, and chromatin structure of and gene-modification experiments with deoxyribonucleic acid (DNA) are described. Indicates that DNA's double helix is variable and also flexible as it interacts with regulatory and other molecules to transfer hereditary messages. (DH)

  16. Silver Nanoscale Hexagonal Column Chips for Detecting Cell-free DNA and Circulating Nucleosomes in Cancer Patients

    PubMed Central

    Ito, Hiroaki; Hasegawa, Katsuyuki; Hasegawa, Yuuki; Nishimaki, Tadashi; Hosomichi, Kazuyoshi; Kimura, Satoshi; Ohba, Motoi; Yao, Hiroshi; Onimaru, Manabu; Inoue, Ituro; Inoue, Haruhiro

    2015-01-01

    Blood tests, which are commonly used for cancer screening, generally have low sensitivity. Here, we developed a novel rapid and simple method to generate silver nanoscale hexagonal columns (NHCs) for use in surface-enhanced Raman scattering (SERS). We reported that the intensity of SERS spectra of clinical serum samples obtained from gastrointestinal cancer patients is was significantly higher than that of SERS spectra of clinical serum samples obtained from non-cancer patients. We estimated the combined constituents on silver NHCs by using a field emission-type scanning electron microscope, Raman microscopes, and a 3D laser scanning confocal microscope. We obtained the Raman scattering spectra of samples of physically fractured cells and clinical serum. No spectra were obtained for chemically lysed cultured cells and DNA, RNA, and protein extracted from cultured cells. We believe that our method, which uses SERS with silver NHCs to detect circulating nucleosomes bound by methylated cell-free DNA, may be successfully implemented in blood tests for cancer screening. PMID:25994878

  17. Lab-on-a-chip-based PCR-RFLP assay for the confirmed detection of short-length feline DNA in food.

    PubMed

    Ali, Md Eaqub; Al Amin, Md; Hamid, Sharifah Bee Abd; Hossain, M A Motalib; Mustafa, Shuhaimi

    2015-01-01

    Wider availability but lack of legal market trades has given feline meat a high potential for use as an adulterant in common meat and meat products. However, mixing of feline meat or its derivatives in food is a sensitive issue, since it is a taboo in most countries and prohibited in certain religions such as Islam and Judaism. Cat meat also has potential for contamination with of severe acute respiratory syndrome, anthrax and hepatitis, and its consumption might lead to an allergic reaction. We developed a very short-amplicon-length (69 bp) PCR assay, authenticated the amplified PCR products by AluI-restriction digestion followed by its separation and detection on a lab-on-a-chip-based automated electrophoretic system, and proved its superiority over the existing long-amplicon-based assays. Although it has been assumed that longer DNA targets are susceptible to breakdown under compromised states, scientific evidence for this hypothesis has been rarely documented. Strong evidence showed that shorter targets are more stable than the longer ones. We confirmed feline-specificity by cross-challenging the primers against 10 different species of terrestrial, aquatic and plant origins in the presence of a 141-bp site of an 18S rRNA gene as a universal eukaryotic control. RFLP analysis separated 43- and 26-bp fragments of AluI-digest in both the gel-image and electropherograms, confirming the original products. The tested detection limit was 0.01% (w/w) feline meat in binary and ternary admixed as well as meatball matrices. Shorter target, better stability and higher sensitivity mean such an assay would be valid for feline identification even in degraded specimens. PMID:26208950

  18. Solid-phase based on-chip DNA purification through a valve-free stepwise injection of multiple reagents employing centrifugal force combined with a hydrophobic capillary barrier pressure.

    PubMed

    Zhang, Hainan; Tran, Hong Hanh; Chung, Bong Hyun; Lee, Nae Yoon

    2013-03-21

    In this paper, we demonstrate a simple technique for sequentially introducing multiple sample liquids into microchannels driven by centrifugal force combined with a hydrophobic barrier pressure and apply the technique for performing solid-phase based on-chip DNA purification. Three microchannels with varying widths, all equipped with independent sample reservoirs at the inlets, were fabricated on a hydrophobic elastomer, poly(dimethylsiloxane) (PDMS). First, glass beads were packed inside the reaction chamber, and a whole cell containing the DNA extract was introduced into the widest channel by applying centrifugal force for physical adsorption of the DNA onto the glass beads. Next, washing and elution solutions were sequentially introduced into the intermediate and narrowest microchannels, respectively, by gradually increasing the amount of centrifugal force. Through a precise manipulation of the centrifugal force, the DNA adsorbed onto the glass beads was successfully washed and eluted in a continuous manner without the need to introduce each solution manually. A stepwise injection of liquids was successfully demonstrated using multiple ink solutions, the results of which corresponded well with the theoretical analyses. As a practical application, the D1S80 locus of human genomic DNA, which is widely used for forensic purposes, was successfully purified using the microdevice introduced in this study, as demonstrated through successful target amplification. This will pave the way for the construction of a control-free valve system for realizing on-chip DNA purification, which is one of the most labor-intensive and hard-to-miniaturize components, on a greatly simplified and miniaturized platform employing hydrophobic PDMS.

  19. Exploring Genome-wide DNA Methylation Profiles Altered in Kashin-Beck Disease Using Infinium Human Methylation 450 Bead Chips.

    PubMed

    Shi, Xiao Wei; Shi, Bo Hui; Lyu, Ai Li; Zhang, Feng; Zhou, Tian Tian; Guo, Xiong

    2016-07-01

    To understand how differentially methylated genes (DMGs) might affect the pathogenesis of Kashin-Beck disease (KBD). Genome-wide methylation profiling of whole blood from 12 matched KBD and controls pairs was performed using a high-resolution Infinium 450 K methylation array. In total, 97 CpG sites were differentially methylated in KBD compared to the normal controls; of these sites, 36 sites were significantly hypermethylated (covering 22 genes) and 61 sites were significantly hypomethylated (covering 34 genes). Of these genes, 14 significant pathways were identified, the most significant P value pathway was type I diabetes mellitus pathway and pathways associated with autoimmune diseases and inflammatory diseases were included in this study. Subsequently, 4 CpG sites in HLA-DRB1 were validated using bisulfite sequencing polymerase chain reaction (BSP) in articular cartilage, and the results showed significant differences in the methylation status between KBD and controls, consistent with the results of the high-resolution array. These results suggested that differences in genome-wide DNA methylation exist between KBD and the controls, and the biological pathways support the autoimmune disease and inflammatory disease hypothesis of KBD.

  20. Exploring Genome-wide DNA Methylation Profiles Altered in Kashin-Beck Disease Using Infinium Human Methylation 450 Bead Chips.

    PubMed

    Shi, Xiao Wei; Shi, Bo Hui; Lyu, Ai Li; Zhang, Feng; Zhou, Tian Tian; Guo, Xiong

    2016-07-01

    To understand how differentially methylated genes (DMGs) might affect the pathogenesis of Kashin-Beck disease (KBD). Genome-wide methylation profiling of whole blood from 12 matched KBD and controls pairs was performed using a high-resolution Infinium 450 K methylation array. In total, 97 CpG sites were differentially methylated in KBD compared to the normal controls; of these sites, 36 sites were significantly hypermethylated (covering 22 genes) and 61 sites were significantly hypomethylated (covering 34 genes). Of these genes, 14 significant pathways were identified, the most significant P value pathway was type I diabetes mellitus pathway and pathways associated with autoimmune diseases and inflammatory diseases were included in this study. Subsequently, 4 CpG sites in HLA-DRB1 were validated using bisulfite sequencing polymerase chain reaction (BSP) in articular cartilage, and the results showed significant differences in the methylation status between KBD and controls, consistent with the results of the high-resolution array. These results suggested that differences in genome-wide DNA methylation exist between KBD and the controls, and the biological pathways support the autoimmune disease and inflammatory disease hypothesis of KBD. PMID:27554126

  1. Immunoassay, DNA Analysis, and Other Ligand Binding Assay Techniques: From Electropherograms to Multiplexed, Ultrasensitive Microarrays on a Chip

    NASA Astrophysics Data System (ADS)

    Ekins, Roger P.

    1999-06-01

    "Ligand" or "binding" assays have made a major impact on biomedical research and clinical diagnosis since their development in the late 1950s. Immunoassay techniques (relying on specific antibodies to bind the target analyte) represent the best-known example, but analogous DNA and RNA analysis methods (using oligonucleotides to recognize defined polynucleotide sequences) are rapidly gaining in importance and are likely to exert profound effects on human society. The evolution of these methods may be divided into three phases: (i) the initial development and widespread use of sensitive "competitive" assays relying on radioisotopically labeled analyte to monitor the binding reaction; (ii) the introduction in the 1980s of "ultrasensitive", "noncompetitive", labeled antibody methods relying on high-specific-activity nonisotopic labels, leading to the emergence of the automatic analyzers that now dominate the field, and (iii) the present development of "microarray"methods based on antibody or oligonucleotide microspots (each recognizing an individual analyte) arrayed on a solid support and relying on observation (typically by confocal microscopy) of fluorescent signals emitted from each spot. Miniaturized microarray methods permitting ultrasensitive measurement of hundreds of different analytes in a minute sample are likely to revolutionize medicine and related fields within the next decade.

  2. Analysis of gene repair tracts from Cas9/gRNA double-stranded breaks in the human CFTR gene

    PubMed Central

    Hollywood, Jennifer A.; Lee, Ciaran M.; Scallan, Martina F.; Harrison, Patrick T.

    2016-01-01

    To maximise the efficiency of template-dependent gene editing, most studies describe programmable and/or RNA-guided endonucleases that make a double-stranded break at, or close to, the target sequence to be modified. The rationale for this design strategy is that most gene repair tracts will be very short. Here, we describe a CRISPR Cas9/gRNA selection-free strategy which uses deep sequencing to characterise repair tracts from a donor plasmid containing seven nucleotide differences across a 216 bp target region in the human CFTR gene. We found that 90% of the template-dependent repair tracts were >100 bp in length with equal numbers of uni-directional and bi-directional repair tracts. The occurrence of long repair tracts suggests that a single gRNA could be used with variants of the same template to create or correct specific mutations within a 200 bp range, the size of ~80% of human exons. The selection-free strategy used here also allowed detection of non-homologous end joining events in many of the homology-directed repair tracts. This indicates a need to modify the donor, possibly by silent changes in the PAM sequence, to prevent creation of a second double-stranded break in an allele that has already been correctly edited by homology-directed repair. PMID:27557525

  3. Analysis of gene repair tracts from Cas9/gRNA double-stranded breaks in the human CFTR gene.

    PubMed

    Hollywood, Jennifer A; Lee, Ciaran M; Scallan, Martina F; Harrison, Patrick T

    2016-01-01

    To maximise the efficiency of template-dependent gene editing, most studies describe programmable and/or RNA-guided endonucleases that make a double-stranded break at, or close to, the target sequence to be modified. The rationale for this design strategy is that most gene repair tracts will be very short. Here, we describe a CRISPR Cas9/gRNA selection-free strategy which uses deep sequencing to characterise repair tracts from a donor plasmid containing seven nucleotide differences across a 216 bp target region in the human CFTR gene. We found that 90% of the template-dependent repair tracts were >100 bp in length with equal numbers of uni-directional and bi-directional repair tracts. The occurrence of long repair tracts suggests that a single gRNA could be used with variants of the same template to create or correct specific mutations within a 200 bp range, the size of ~80% of human exons. The selection-free strategy used here also allowed detection of non-homologous end joining events in many of the homology-directed repair tracts. This indicates a need to modify the donor, possibly by silent changes in the PAM sequence, to prevent creation of a second double-stranded break in an allele that has already been correctly edited by homology-directed repair. PMID:27557525

  4. DNA

    ERIC Educational Resources Information Center

    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)

  5. Design and experiment of silicon PCR chips

    NASA Astrophysics Data System (ADS)

    Cui, Zheng; Zhao, Zhan; Xia, Shanhong

    2002-04-01

    There are considerable interests in integrating Polymerase chain reaction (PCR) on a microchip can have much fast heating and cooling rate, the delicacy in its structure makes the PCR experiment difficult and cracks often occur particularly for the thin membrane type of PCR chips. Design study and experiment of silicon PCR chips are presented with the aim of identifying the problems encountered in experiment and finding an optimum chip structure. Heating characteristics of four different heater designs have been compared, so have the PCR chambers with fixed frame and with suspended frame. The thermal stress analysis has shown that the structure and heater design can make a significant difference in heating characteristics and in reducing the failure of PCR chips. Different solutions to reduce PCR chip failure have been proposed. One of the solutions was implemented in the experiment, confirming the design study results. Silicon PCR chips have been fabricated. Thermal cycling and initial DNA amplification results are presented.

  6. The shuttling SR protein 9G8 plays a role in translation of unspliced mRNA containing a constitutive transport element.

    PubMed

    Swartz, Jennifer E; Bor, Yeou-Cherng; Misawa, Yukiko; Rekosh, David; Hammarskjold, Marie-Louise

    2007-07-01

    The splicing regulatory SR protein, 9G8, has recently been proposed to function in mRNA export in conjunction with the export protein, Tap/NXF1. Tap interacts directly with the Mason-Pfizer monkey virus constitutive transport element (CTE), an element that enables export of unspliced, intron-containing mRNA. Based on our previous finding that Tap can promote polysome association and translation of CTE-RNA, we investigated the effect of 9G8 on cytoplasmic RNA fate. 9G8 was shown to enhance expression of unspliced RNA containing either the Mason-Pfizer monkey virus-CTE or the recently discovered Tap-CTE. 9G8 also enhanced polyribosome association of unspliced RNA containing a CTE. Hyperphosphorylated 9G8 was present in monosomes and small polyribosomes, whereas soluble fractions contained only hypophosphorylated protein. Our results are consistent with a model in which hypophosphorylated SR proteins remain stably associated with messenger ribonucleoprotein (mRNP) complexes during export and are released during translation initiation concomitant with increased phosphorylation. These results provide further evidence for crucial links between RNA splicing, export and translation. PMID:17513303

  7. The characterization of four gene expression analysis in circulating tumor cells made by Multiplex-PCR from the AdnaTest kit on the lab-on-a-chip Agilent DNA 1000 platform

    PubMed Central

    Škereňová, Markéta; Mikulová, Veronika; Čapoun, Otakar; Zima, Tomáš

    2016-01-01

    Introduction Nowadays, on-a-chip capillary electrophoresis is a routine method for the detection of PCR fragments. The Agilent 2100 Bioanalyzer was one of the first commercial devices in this field. Our project was designed to study the characteristics of Agilent DNA 1000 kit in PCR fragment analysis as a part of circulating tumour cell (CTC) detection technique. Despite the common use of this kit a complex analysis of the results from a long-term project is still missing. Materials and methods A commercially available Agilent DNA 1000 kit was used as a final step in the CTC detection (AdnaTest) for the determination of the presence of PCR fragments generated by Multiplex PCR. Data from 30 prostate cancer patients obtained during two years of research were analyzed to determine the trueness and precision of the PCR fragment size determination. Additional experiments were performed to demonstrate the precision (repeatability, reproducibility) and robustness of PCR fragment concentration determination. Results The trueness and precision of the size determination was below 3% and 2% respectively. The repeatability of the concentration determination was below 15%. The difference in concentration determination increases when Multiplex-PCR/storage step is added between the two measurements of one sample. Conclusions The characteristics established in our study are in concordance with the manufacturer’s specifications established for a ladder as a sample. However, the concentration determination may vary depending on chip preparation, sample storage and concentration. The 15% variation of concentration determination repeatability was shown to be partly proportional and can be suppressed by proper normalization. PMID:26981024

  8. Multicolor and Erasable DNA Photolithography

    PubMed Central

    2015-01-01

    The immobilization of DNA molecules onto a solid support is a crucial step in biochip research and related applications. In this work, we report a DNA photolithography method based on photocleavage of 2-nitrobenzyl linker-modified DNA strands. These strands were subjected to ultraviolet light irradiation to generate multiple short DNA strands in a programmable manner. Coupling the toehold-mediated DNA strand-displacement reaction with DNA photolithography enabled the fabrication of a DNA chip surface with multifunctional DNA patterns having complex geometrical structures at the microscale level. The erasable DNA photolithography strategy was developed to allow different paintings on the same chip. Furthermore, the asymmetrical modification of colloidal particles was carried out by using this photolithography strategy. This strategy has broad applications in biosensors, nanodevices, and DNA-nanostructure fabrication. PMID:24988147

  9. Multicolor and erasable DNA photolithography.

    PubMed

    Huang, Fujian; Xu, Huaguo; Tan, Weihong; Liang, Haojun

    2014-07-22

    The immobilization of DNA molecules onto a solid support is a crucial step in biochip research and related applications. In this work, we report a DNA photolithography method based on photocleavage of 2-nitrobenzyl linker-modified DNA strands. These strands were subjected to ultraviolet light irradiation to generate multiple short DNA strands in a programmable manner. Coupling the toehold-mediated DNA strand-displacement reaction with DNA photolithography enabled the fabrication of a DNA chip surface with multifunctional DNA patterns having complex geometrical structures at the microscale level. The erasable DNA photolithography strategy was developed to allow different paintings on the same chip. Furthermore, the asymmetrical modification of colloidal particles was carried out by using this photolithography strategy. This strategy has broad applications in biosensors, nanodevices, and DNA-nanostructure fabrication.

  10. Lab-on-a-chip technologies for single-molecule studies.

    PubMed

    Zhao, Yanhui; Chen, Danqi; Yue, Hongjun; French, Jarrod B; Rufo, Joseph; Benkovic, Stephen J; Huang, Tony Jun

    2013-06-21

    Recent developments on various lab-on-a-chip techniques allow miniaturized and integrated devices to perform on-chip single-molecule studies. Fluidic-based platforms that utilize unique microscale fluidic behavior are capable of conducting single-molecule experiments with high sensitivities and throughputs, while biomolecular systems can be studied on-chip using techniques such as DNA curtains, magnetic tweezers, and solid-state nanopores. The advances of these on-chip single-molecule techniques lead to next-generation lab-on-a-chip devices, such as DNA transistors, and single-molecule real-time (SMRT) technology for rapid and low-cost whole genome DNA sequencing. In this Focus article, we will discuss some recent successes in the development of lab-on-a-chip techniques for single-molecule studies and expound our thoughts on the near future of on-chip single-molecule studies.

  11. Lab-on-a-chip technologies for single-molecule studies

    PubMed Central

    Zhao, Yanhui; Chen, Danqi; Yue, Hongjun; French, Jarrod B.; Rufo, Joey; Benkovic, Stephen J.; Huang, Tony Jun

    2014-01-01

    Recent developments on various lab-on-a-chip techniques allow miniaturized and integrated devices to perform on-chip single-molecule studies. Fluidic-based platforms that utilize the unique microscale fluidic behavior are capable of conducting single-molecule experiments with high sensitivities and throughputs, while biomolecular systems can be studied on-chip using techniques such as DNA curtains, magnetic tweezers, and solid-state nanopores. The advances of these on-chip single-molecule techniques lead to next-generation lab-on-a-chip devices such as DNA transistors, and single-molecule real-time (SMRT) technology for rapid and low-cost whole genome DNA sequencing. In this Focus article, we will discuss some recent successes on developing lab-on-a-chip techniques for single-molecule studies and expound our thoughts on the near future of on-chip single-molecule studies. PMID:23670195

  12. Microfabricated integrated DNA analysis systems

    SciTech Connect

    Woolley, A.T.; Mathies, R.A.; Northrup, M.A.

    1996-12-31

    Microfabrication has the potential to revolutionize chemical analysis, from reactions to separations to molecular biotechnology. Microfabricated devices allow high speed separations, automated sample handling, and the study of reactions in the pl to {mu}l volume range. Our research has focused on microfabricated integrated DNA analysis systems (MIDAS). As a first step, we have demonstrated high-speed DNA restriction fragment sizing, and DNA sequencing on microfabricated capillary electrophoresis (CE) chips. We have recently coupled microfabricated PCR reactors and CE chips to make integrated DNA analysis devices. With these devices, rapid PCR can be performed and the reaction products immediately analyzed on the CE chip, eliminating the need for manual transfer of the amplified sample. PCR amplifications have been done in less than 16 minutes, followed by CE analysis in under 100 seconds. These PCR-CE chips represent an important step towards completely integrated sample manipulation on microfabricated devices.

  13. Solvent resistant microfluidic DNA synthesizer.

    PubMed

    Huang, Yanyi; Castrataro, Piero; Lee, Cheng-Chung; Quake, Stephen R

    2007-01-01

    We fabricated a microfluidic DNA synthesizer out of perfluoropolyether (PFPE), an elastomer with excellent chemical compatibility which makes it possible to perform organic chemical reactions, and synthesized 20-mer oligonucleotides on chip. PMID:17180201

  14. Gene Chips and Functional Genomics

    NASA Astrophysics Data System (ADS)

    Hamadeh, Hisham; Afshari, Cynthia

    2000-11-01

    These past few years of scientific discovery will undoubtedly be remembered as the "genomics era," the period in which biologists succeeded in enumerating the sequence of nucleotides making up all, or at least most, of human DNA. And while this achievement has been heralded as a technological feat equal to the moon landing, it is only the first of many advances in DNA technology. Scientists are now faced with the task of understanding the meaning of the DNA sequence. Specifically, they want to learn how the DNA code relates to protein function. An important tool in the study of "functional genomics," is the cDNA microarray—also known as the gene chip. Inspired by computer microchips, gene chips allow scientists to monitor the expression of hundreds, even thousands, of genes in a fraction of the time it used to take to monitor the expression of a single one. By altering the conditions under which a particular tissue expresses genes—say, by exposing it to toxins or growth factors—scientists can determine the suite of genes expressed in different situations and hence start to get a handle on the function of these genes. The authors discuss this important new technology and some of its practical applications.

  15. Resveratrol overcomes gefitinib resistance by increasing the intracellular gefitinib concentration and triggering apoptosis, autophagy and senescence in PC9/G NSCLC cells

    PubMed Central

    Zhu, Yinsong; He, Wenjuan; Gao, Xiujuan; Li, Bin; Mei, Chenghan; Xu, Rong; Chen, Hui

    2015-01-01

    Gefitinib (Gef) provides clinical benefits to non-small cell lung cancer (NSCLC) patients with activating EGFR mutations. However, acquired resistance (AR) is a major obstacle to effective Gef therapy. This study demonstrated that resveratrol (Res) could synergize with Gef to inhibit the proliferation of Gef-resistant NSCLC cells. The underlying mechanisms of synergism were investigated, and the results showed that cotreatment with Gef and Res could inhibit EGFR phosphorylation by increasing intracellular Gef accumulation through the impairment of Gef elimination from PC9/G cells. Consistently, CYP1A1 and ABCG2 expression were inhibited. Meanwhile, the cotreatment significantly induced cell apoptosis, autophagy, cell cycle arrest and senescence accompanied by increased expression of cleaved caspase-3, LC3B-II, p53 and p21. Further studies revealed that autophagy inhibition enhanced apoptosis and abrogated senescence while apoptosis inhibition had no notable effect on cell autophagy and senescence during cotreatment with Gef and Res. These results indicated that in addition to apoptosis, senescence promoted by autophagy contributes to the antiproliferation effect of combined Gef and Res on PC9/G cells. In conclusion, combined treatment with Gef and Res may represent a rational strategy to overcome AR in NSCLC cells. PMID:26635117

  16. Detection and quantitation of single nucleotide polymorphisms, DNA sequence variations, DNA mutations, DNA damage and DNA mismatches

    DOEpatents

    McCutchen-Maloney, Sandra L.

    2002-01-01

    DNA mutation binding proteins alone and as chimeric proteins with nucleases are used with solid supports to detect DNA sequence variations, DNA mutations and single nucleotide polymorphisms. The solid supports may be flow cytometry beads, DNA chips, glass slides or DNA dips sticks. DNA molecules are coupled to solid supports to form DNA-support complexes. Labeled DNA is used with unlabeled DNA mutation binding proteins such at TthMutS to detect DNA sequence variations, DNA mutations and single nucleotide length polymorphisms by binding which gives an increase in signal. Unlabeled DNA is utilized with labeled chimeras to detect DNA sequence variations, DNA mutations and single nucleotide length polymorphisms by nuclease activity of the chimera which gives a decrease in signal.

  17. Development of Microreactor Array Chip-Based Measurement System for Massively Parallel Analysis of Enzymatic Activity

    NASA Astrophysics Data System (ADS)

    Hosoi, Yosuke; Akagi, Takanori; Ichiki, Takanori

    Microarray chip technology such as DNA chips, peptide chips and protein chips is one of the promising approaches for achieving high-throughput screening (HTS) of biomolecule function since it has great advantages in feasibility of automated information processing due to one-to-one indexing between array position and molecular function as well as massively parallel sample analysis as a benefit of down-sizing and large-scale integration. Mostly, however, the function that can be evaluated by such microarray chips is limited to affinity of target molecules. In this paper, we propose a new HTS system of enzymatic activity based on microreactor array chip technology. A prototype of the automated and massively parallel measurement system for fluorometric assay of enzymatic reactions was developed by the combination of microreactor array chips and a highly-sensitive fluorescence microscope. Design strategy of microreactor array chips and an optical measurement platform for the high-throughput enzyme assay are discussed.

  18. Development and application of compact and on-chip electron linear accelerators for dynamic tracking cancer therapy and DNA damage/repair analysis

    NASA Astrophysics Data System (ADS)

    Uesaka, M.; Demachi, K.; Fujiwara, T.; Dobashi, K.; Fujisawa, H.; Chhatkuli, R. B.; Tsuda, A.; Tanaka, S.; Matsumura, Y.; Otsuki, S.; Kusano, J.; Yamamoto, M.; Nakamura, N.; Tanabe, E.; Koyama, K.; Yoshida, M.; Fujimori, R.; Yasui, A.

    2015-06-01

    We are developing compact electron linear accelerators (hereafter linac) with high RF (Radio Frequency) frequency (9.3 GHz, wavelength 32.3 mm) of X-band and applying to medicine and non-destructive testing. Especially, potable 950 keV and 3.95 MeV linac X-ray sources have been developed for on-site transmission testing at several industrial plants and civil infrastructures including bridges. 6 MeV linac have been made for pinpoint X-ray dynamic tracking cancer therapy. The length of the accelerating tube is ∼600 mm. The electron beam size at the X-ray target is less than 1 mm and X-ray spot size at the cancer is less than 3 mm. Several hardware and software are under construction for dynamic tracking therapy for moving lung cancer. Moreover, as an ultimate compact linac, we are designing and manufacturing a laser dielectric linac of ∼1 MeV with Yr fiber laser (283 THz, wavelength 1.06 pm). Since the wavelength is 1.06 μm, the length of one accelerating strcture is tens pm and the electron beam size is in sub-micro meter. Since the sizes of cell and nuclear are about 10 and 1 μm, respectively, we plan to use this “On-chip” linac for radiation-induced DNA damage/repair analysis. We are thinking a system where DNA in a nucleus of cell is hit by ∼1 μm electron or X-ray beam and observe its repair by proteins and enzymes in live cells in-situ.

  19. Integrated chip-based capillary electrophoresis.

    PubMed

    Effenhauser, C S; Bruin, G J; Paulus, A

    1997-11-01

    Integrated capillary electrophoresis (ICE) is emerging as a new analytical tool allowing fast, automated, miniaturized and multiplexed assays, thus meeting the needs of the pharmaceutical industry in drug development. The current state-of-the-art of ICE is described with an emphasis on the choice of the support material (glass or polymeric materials), electrokinetic fluid handling, and injection and detection issues. Strategies and chip designs for pre- or post-column derivatization, DNA sequencing, on-line PCR analysis, on-chip enzymatic sample digestion, fraction isolation, and immunoassays are presented. The review concludes with a brief outlook.

  20. PHYSICS: Toward Atom Chips.

    PubMed

    Fortágh, József; Zimmermann, Claus

    2005-02-11

    As a novel approach for turning the peculiar features of quantum mechanics into practical devices, researchers are investigating the use of ultracold atomic clouds above microchips. Such "atom chips" may find use as sensitive probes for gravity, acceleration, rotation, and tiny magnetic forces. In their Perspective, Fortagh and Zimmermann discuss recent advances toward creating atom chips, in which current-carrying conductors in the chips create magnetic microtraps that confine the atomic clouds. Despite some intrinsic limits to the performance of atom chips, existing technologies are capable of producing atom chips, and many possibilities for their construction remain to be explored.

  1. ChIP analysis unravels an exceptionally wide distribution of DNA binding sites for the NtcA transcription factor in a heterocyst-forming cyanobacterium

    PubMed Central

    2014-01-01

    Background The CRP-family transcription factor NtcA, universally found in cyanobacteria, was initially discovered as a regulator operating N control. It responds to the N regime signaled by the internal 2-oxoglutarate levels, an indicator of the C to N balance of the cells. Canonical NtcA-activated promoters bear an NtcA-consensus binding site (GTAN8TAC) centered at about 41.5 nucleotides upstream from the transcription start point. In strains of the Anabaena/Nostoc genera NtcA is pivotal for the differentiation of heterocysts in response to N stress. Results In this study, we have used chromatin immunoprecipitation followed by high-throughput sequencing to identify the whole catalog of NtcA-binding sites in cells of the filamentous, heterocyst-forming cyanobacterium Anabaena sp. PCC 7120 three hours after the withdrawal of combined N. NtcA has been found to bind to 2,424 DNA regions in the genome of Anabaena, which have been ascribed to 2,153 genes. Interestingly, only a small proportion of those genes are involved in N assimilation and metabolism, and 65% of the binding regions were located intragenically. Conclusions The distribution of NtcA-binding sites identified here reveals the largest bacterial regulon described to date. Our results show that NtcA has a much wider role in the physiology of the cell than it has been previously thought, acting both as a global transcriptional regulator and possibly also as a factor influencing the superstructure of the chromosome (and plasmids). PMID:24417914

  2. The GenoChip: a new tool for genetic anthropology.

    PubMed

    Elhaik, Eran; Greenspan, Elliott; Staats, Sean; Krahn, Thomas; Tyler-Smith, Chris; Xue, Yali; Tofanelli, Sergio; Francalacci, Paolo; Cucca, Francesco; Pagani, Luca; Jin, Li; Li, Hui; Schurr, Theodore G; Greenspan, Bennett; Spencer Wells, R

    2013-01-01

    The Genographic Project is an international effort aimed at charting human migratory history. The project is nonprofit and nonmedical, and, through its Legacy Fund, supports locally led efforts to preserve indigenous and traditional cultures. Although the first phase of the project was focused on uniparentally inherited markers on the Y-chromosome and mitochondrial DNA (mtDNA), the current phase focuses on markers from across the entire genome to obtain a more complete understanding of human genetic variation. Although many commercial arrays exist for genome-wide single-nucleotide polymorphism (SNP) genotyping, they were designed for medical genetic studies and contain medically related markers that are inappropriate for global population genetic studies. GenoChip, the Genographic Project's new genotyping array, was designed to resolve these issues and enable higher resolution research into outstanding questions in genetic anthropology. The GenoChip includes ancestry informative markers obtained for over 450 human populations, an ancient human (Saqqaq), and two archaic hominins (Neanderthal and Denisovan) and was designed to identify all known Y-chromosome and mtDNA haplogroups. The chip was carefully vetted to avoid inclusion of medically relevant markers. To demonstrate its capabilities, we compared the FST distributions of GenoChip SNPs to those of two commercial arrays. Although all arrays yielded similarly shaped (inverse J) FST distributions, the GenoChip autosomal and X-chromosomal distributions had the highest mean FST, attesting to its ability to discern subpopulations. The chip performances are illustrated in a principal component analysis for 14 worldwide populations. In summary, the GenoChip is a dedicated genotyping platform for genetic anthropology. With an unprecedented number of approximately 12,000 Y-chromosomal and approximately 3,300 mtDNA SNPs and over 130,000 autosomal and X-chromosomal SNPs without any known health, medical, or phenotypic

  3. The GenoChip: A New Tool for Genetic Anthropology

    PubMed Central

    Elhaik, Eran; Greenspan, Elliott; Staats, Sean; Krahn, Thomas; Tyler-Smith, Chris; Xue, Yali; Tofanelli, Sergio; Francalacci, Paolo; Cucca, Francesco; Pagani, Luca; Jin, Li; Li, Hui; Schurr, Theodore G.; Greenspan, Bennett; Spencer Wells, R.

    2013-01-01

    The Genographic Project is an international effort aimed at charting human migratory history. The project is nonprofit and nonmedical, and, through its Legacy Fund, supports locally led efforts to preserve indigenous and traditional cultures. Although the first phase of the project was focused on uniparentally inherited markers on the Y-chromosome and mitochondrial DNA (mtDNA), the current phase focuses on markers from across the entire genome to obtain a more complete understanding of human genetic variation. Although many commercial arrays exist for genome-wide single-nucleotide polymorphism (SNP) genotyping, they were designed for medical genetic studies and contain medically related markers that are inappropriate for global population genetic studies. GenoChip, the Genographic Project’s new genotyping array, was designed to resolve these issues and enable higher resolution research into outstanding questions in genetic anthropology. The GenoChip includes ancestry informative markers obtained for over 450 human populations, an ancient human (Saqqaq), and two archaic hominins (Neanderthal and Denisovan) and was designed to identify all known Y-chromosome and mtDNA haplogroups. The chip was carefully vetted to avoid inclusion of medically relevant markers. To demonstrate its capabilities, we compared the FST distributions of GenoChip SNPs to those of two commercial arrays. Although all arrays yielded similarly shaped (inverse J) FST distributions, the GenoChip autosomal and X-chromosomal distributions had the highest mean FST, attesting to its ability to discern subpopulations. The chip performances are illustrated in a principal component analysis for 14 worldwide populations. In summary, the GenoChip is a dedicated genotyping platform for genetic anthropology. With an unprecedented number of approximately 12,000 Y-chromosomal and approximately 3,300 mtDNA SNPs and over 130,000 autosomal and X-chromosomal SNPs without any known health, medical, or phenotypic

  4. DNA Microarrays

    NASA Astrophysics Data System (ADS)

    Nguyen, C.; Gidrol, X.

    Genomics has revolutionised biological and biomedical research. This revolution was predictable on the basis of its two driving forces: the ever increasing availability of genome sequences and the development of new technology able to exploit them. Up until now, technical limitations meant that molecular biology could only analyse one or two parameters per experiment, providing relatively little information compared with the great complexity of the systems under investigation. This gene by gene approach is inadequate to understand biological systems containing several thousand genes. It is essential to have an overall view of the DNA, RNA, and relevant proteins. A simple inventory of the genome is not sufficient to understand the functions of the genes, or indeed the way that cells and organisms work. For this purpose, functional studies based on whole genomes are needed. Among these new large-scale methods of molecular analysis, DNA microarrays provide a way of studying the genome and the transcriptome. The idea of integrating a large amount of data derived from a support with very small area has led biologists to call these chips, borrowing the term from the microelectronics industry. At the beginning of the 1990s, the development of DNA chips on nylon membranes [1, 2], then on glass [3] and silicon [4] supports, made it possible for the first time to carry out simultaneous measurements of the equilibrium concentration of all the messenger RNA (mRNA) or transcribed RNA in a cell. These microarrays offer a wide range of applications, in both fundamental and clinical research, providing a method for genome-wide characterisation of changes occurring within a cell or tissue, as for example in polymorphism studies, detection of mutations, and quantitative assays of gene copies. With regard to the transcriptome, it provides a way of characterising differentially expressed genes, profiling given biological states, and identifying regulatory channels.

  5. Causes of stem end chip defect in chipping potatoes

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Stem-end chip defect (SECD) is a serious tuber quality concern that affects chipping potatoes. This defect is characterized by dark-colored vascular tissues and adjacent cortical tissues at the tuber stem-end of potato chips after frying. Chips with SECD are unappealing to consumers and raw product ...

  6. Novel, rapid DNA-based on-chip bacterial identification system combining dielectrophoresis and amplification-free fluorescent resonance energy transfer assisted in-situ hybridization (FRET-ISH)

    NASA Astrophysics Data System (ADS)

    Packard, Michelle M.; Shusteff, Maxim; Alocilja, Evangelyn

    2011-10-01

    Although real-time PCR (RT-PCR) has become a diagnostic standard for rapid identification of bacterial species, typical methods remain time-intensive due to sample preparation and amplification cycle times. The assay described in this work incorporates on-chip dielectrophoretic capture and concentration of bacterial cells, thermal lysis, cell permeabilization, and nucleic acid denaturation and fluorescence resonance energy transfer assisted in-situ hybridization (FRET-ISH) species identification. Identification is achieved completely on chip in less than thirty minutes from receipt of sample compared to multiple hours required by traditional RT-PCR and its requisite sample preparation.

  7. CHIP, CHIP, ARRAY! THREE CHIPS FOR POST-GENOMIC RESEARCH

    EPA Science Inventory

    Cambridge Healthtech Institute recently held the 4th installment of their popular "Lab-on-a-Chip" series in Zurich, Switzerland. As usual, it was enthusiastically received and over 225 people attended the 2-1/2 day meeting to see and hear about some of the latest developments an...

  8. Carbonyl sulfide removal with compost and wood chip biofilters, and in the presence of hydrogen sulfide.

    PubMed

    Sattler, Melanie L; Garrepalli, Divya R; Nawal, Chandraprakash S

    2009-12-01

    Carbonyl sulfide (COS) is an odor-causing compound and hazardous air pollutant emitted frequently from wastewater treatment facilities and chemical and primary metals industries. This study examined the effectiveness of biofiltration in removing COS. Specific objectives were to compare COS removal efficiency for various biofilter media; to determine whether hydrogen sulfide (H2S), which is frequently produced along with COS under anaerobic conditions, adversely impacts COS removal; and to determine the maximum elimination capacity of COS for use in biofilter design. Three laboratory-scale polyvinyl chloride biofilter columns were filled with up to 28 in. of biofilter media (aged compost, fresh compost, wood chips, or a compost/wood chip mixture). Inlet COS ranged from 5 to 46 parts per million (ppm) (0.10-9.0 g/m3 hr). Compost and the compost/wood chip mixture produced higher COS removal efficiencies than wood chips alone. The compost and compost/wood chip mixture had a shorter stabilization times compared with wood chips alone. Fresh versus aged compost did not impact COS removal efficiency. The presence of H2S did not adversely impact COS removal for the concentration ratios tested. The maximum elimination capacity is at least 9 g/m3 hr for COS with compost media.

  9. Carbonyl sulfide removal with compost and wood chip biofilters, and in the presence of hydrogen sulfide.

    PubMed

    Sattler, Melanie L; Garrepalli, Divya R; Nawal, Chandraprakash S

    2009-12-01

    Carbonyl sulfide (COS) is an odor-causing compound and hazardous air pollutant emitted frequently from wastewater treatment facilities and chemical and primary metals industries. This study examined the effectiveness of biofiltration in removing COS. Specific objectives were to compare COS removal efficiency for various biofilter media; to determine whether hydrogen sulfide (H2S), which is frequently produced along with COS under anaerobic conditions, adversely impacts COS removal; and to determine the maximum elimination capacity of COS for use in biofilter design. Three laboratory-scale polyvinyl chloride biofilter columns were filled with up to 28 in. of biofilter media (aged compost, fresh compost, wood chips, or a compost/wood chip mixture). Inlet COS ranged from 5 to 46 parts per million (ppm) (0.10-9.0 g/m3 hr). Compost and the compost/wood chip mixture produced higher COS removal efficiencies than wood chips alone. The compost and compost/wood chip mixture had a shorter stabilization times compared with wood chips alone. Fresh versus aged compost did not impact COS removal efficiency. The presence of H2S did not adversely impact COS removal for the concentration ratios tested. The maximum elimination capacity is at least 9 g/m3 hr for COS with compost media. PMID:20066911

  10. Immobilization of DNA in polyacrylamide gel for the manufacture of DNA and DNA-oligonucleotide microchips.

    SciTech Connect

    Proudnikov, D.; Timofeev, E.; Mirzabekov, A.; Center for Mechanistic Biology and Biotechnology; Engelhardt Inst. of Molecular Biology

    1998-05-15

    Activated DNA was immobilized in aldehyde-containing polyacrylamide gel for use in manufacturing the MAGIChip (microarrays of gel-immobilized compounds on a chip). First, abasic sites were generated in DNA by partial acidic depurination. Amino groups were then introduced into the abasic sites by reaction with ethylenediamine and reduction of the aldimine bonds formed. It was found that DNA could be fragmented at the site of amino group incorporation or preserved mostly unfragmented. In similar reactions, both amino-DNA and amino-oligonucleotides were attached through their amines to polyacrylamide gel derivatized with aldehyde groups. Single- and double-stranded DNA of 40 to 972 nucleotides or base pairs were immobilized on the gel pads to manufacture a DNA microchip. The microchip was hybridized with fluorescently labeled DNA-specific oligonucleotide probes. This procedure for immobilization of amino compounds was used to manufacture MAGIChips containing both DNA and oligonucleotides.

  11. High-Throughput DNA Array for SNP Detection of KRAS Gene Using a Centrifugal Microfluidic Device.

    PubMed

    Sedighi, Abootaleb; Li, Paul C H

    2016-01-01

    Here, we describe detection of single nucleotide polymorphism (SNP) in genomic DNA samples using a NanoBioArray (NBA) chip. Fast DNA hybridization is achieved in the chip when target DNAs are introduced to the surface-arrayed probes using centrifugal force. Gold nanoparticles (AuNPs) are used to assist SNP detection at room temperature. The parallel setting of sample introduction in the spiral channels of the NBA chip enables multiple analyses on many samples, resulting in a technique appropriate for high-throughput SNP detection. The experimental procedure, including chip fabrication, probe array printing, DNA amplification, hybridization, signal detection, and data analysis, is described in detail.

  12. Droplet-based micro oscillating-flow PCR chip

    NASA Astrophysics Data System (ADS)

    Wang, Wei; Li, Zhi-Xin; Luo, Rong; Lü, Shu-Hai; Xu, Ai-Dong; Yang, Yong-Jun

    2005-08-01

    Polymerase chain reactions (PCR), thermally activated chemical reactions which are widely used for nucleic acid amplification, have recently received much attention in microelectromechanical systems and micro total analysis systems because a wide variety of DNA/RNA molecules can be enriched by PCR for further analyses. In the present work, a droplet-based micro oscillating-flow PCR chip was designed and fabricated by the silicon microfabrication technique. Three different temperature zones, which were stable at denaturation, extension and annealing temperatures and isolated from each other by a thin-wall linkage, were integrated with a single, simple and straight microchannel to form the chip's basic functional structure. The PCR mixture was injected into the chip as a single droplet and flowed through the three temperature zones in the main microchannel in an oscillating manner to achieve the temperature maintenance and transitions. The chip's thermal performance was theoretically analyzed and numerically simulated. The results indicated that the time needed for the temperature of the droplet to change to the target value is less than 1 s, and the root mean square error of temperature is less than 0.2 °C. A droplet of 1 µl PCR mixture with standard HPV (Human Papilloma Virus)-DNA sample inside was amplified by the present chip and the results were analyzed by slab gel electrophoresis with separation of DNA markers in parallel. The electrophoresis results demonstrated that the micro oscillating-flow PCR chip successfully amplified the HPV-DNA, with a processing time of about 15 min which is significantly reduced compared to that for the conventional PCR instrument.

  13. Accelerator on a Chip

    ScienceCinema

    England, Joel

    2016-07-12

    SLAC's Joel England explains how the same fabrication techniques used for silicon computer microchips allowed their team to create the new laser-driven particle accelerator chips. (SLAC Multimedia Communications)

  14. Accelerator on a Chip

    SciTech Connect

    England, Joel

    2014-06-30

    SLAC's Joel England explains how the same fabrication techniques used for silicon computer microchips allowed their team to create the new laser-driven particle accelerator chips. (SLAC Multimedia Communications)

  15. Arrays of nucleic acid probes on biological chips

    DOEpatents

    Chee, Mark; Cronin, Maureen T.; Fodor, Stephen P. A.; Huang, Xiaohua X.; Hubbell, Earl A.; Lipshutz, Robert J.; Lobban, Peter E.; Morris, MacDonald S.; Sheldon, Edward L.

    1998-11-17

    DNA chips containing arrays of oligonucleotide probes can be used to determine whether a target nucleic acid has a nucleotide sequence identical to or different from a specific reference sequence. The array of probes comprises probes exactly complementary to the reference sequence, as well as probes that differ by one or more bases from the exactly complementary probes.

  16. ChIP on chip and ChIP-Seq assays: genome-wide analysis of transcription factor binding and histone modifications.

    PubMed

    Pillai, Smitha; Chellappan, Srikumar P

    2015-01-01

    Deregulation of transcriptional activity of many genes has been causatively linked to human diseases including cancer. Altered patterns of gene expression in normal and cancer cells are the result of inappropriate expression of transcription factors and chromatin modifying proteins. Chromatin immunoprecipitation assay is a well-established tool for investigating the interactions between regulatory proteins and DNA at distinct stages of gene activation. ChIP coupled with DNA microarrays, known as ChIP on chip, or sequencing of DNA associated with the factors (ChIP-Seq) allow us to determine the entire spectrum of in vivo DNA binding sites for a given protein. This has been of immense value because ChIP on chip assays and ChIP-Seq experiments can provide a snapshot of the transcriptional regulatory mechanisms on a genome-wide scale. This chapter outlines the general strategies used to carry out ChIP-chip assays to study the differential recruitment of regulatory molecules based on the studies conducted in our lab as well as other published protocols; these can be easily modified to a ChIP-Seq analysis.

  17. Chipping citrus wood for gasifiction

    SciTech Connect

    Churchill, D.B.; Hedden, S.L.; Whitney, J.D.; Shaw, L.N.

    1984-01-01

    Both green and dead citrus trees were used for chipping. Chip moisture content, fuel analysis, drying time, and data on fuel/tonne of chips were obtained. The average moisture contents of green and dead trees when chipped were 25% and 16% (wet basis) respectively. Chips were sized to a minimum of 0.32 squared cm x 0.32 cm thick to a maximum of 5.0 cm squared x 0.32 cm thick and normally required 4 weeks to air dry to 14% (wet basis) moisture content before use. Approximately 50% of the total tree by weight could be made into usable chips. 9 references.

  18. GCOD - GeneChip Oncology Database

    PubMed Central

    2011-01-01

    Background DNA microarrays have become a nearly ubiquitous tool for the study of human disease, and nowhere is this more true than in cancer. With hundreds of studies and thousands of expression profiles representing the majority of human cancers completed and in public databases, the challenge has been effectively accessing and using this wealth of data. Description To address this issue we have collected published human cancer gene expression datasets generated on the Affymetrix GeneChip platform, and carefully annotated those studies with a focus on providing accurate sample annotation. To facilitate comparison between datasets, we implemented a consistent data normalization and transformation protocol and then applied stringent quality control procedures to flag low-quality assays. Conclusion The resulting resource, the GeneChip Oncology Database, is available through a publicly accessible website that provides several query options and analytical tools through an intuitive interface. PMID:21291543

  19. Chip packaging technique

    NASA Technical Reports Server (NTRS)

    Jayaraj, Kumaraswamy (Inventor); Noll, Thomas E. (Inventor); Lockwood, Harry F. (Inventor)

    2001-01-01

    A hermetically sealed package for at least one semiconductor chip is provided which is formed of a substrate having electrical interconnects thereon to which the semiconductor chips are selectively bonded, and a lid which preferably functions as a heat sink, with a hermetic seal being formed around the chips between the substrate and the heat sink. The substrate is either formed of or includes a layer of a thermoplastic material having low moisture permeability which material is preferably a liquid crystal polymer (LCP) and is a multiaxially oriented LCP material for preferred embodiments. Where the lid is a heat sink, the heat sink is formed of a material having high thermal conductivity and preferably a coefficient of thermal expansion which substantially matches that of the chip. A hermetic bond is formed between the side of each chip opposite that connected to the substrate and the heat sink. The thermal bond between the substrate and the lid/heat sink may be a pinched seal or may be provided, for example by an LCP frame which is hermetically bonded or sealed on one side to the substrate and on the other side to the lid/heat sink. The chips may operate in the RF or microwave bands with suitable interconnects on the substrate and the chips may also include optical components with optical fibers being sealed into the substrate and aligned with corresponding optical components to transmit light in at least one direction. A plurality of packages may be physically and electrically connected together in a stack to form a 3D array.

  20. Progress in the application of DNA microarrays.

    PubMed Central

    Lobenhofer, E K; Bushel, P R; Afshari, C A; Hamadeh, H K

    2001-01-01

    Microarray technology has been applied to a variety of different fields to address fundamental research questions. The use of microarrays, or DNA chips, to study the gene expression profiles of biologic samples began in 1995. Since that time, the fundamental concepts behind the chip, the technology required for making and using these chips, and the multitude of statistical tools for analyzing the data have been extensively reviewed. For this reason, the focus of this review will be not on the technology itself but on the application of microarrays as a research tool and the future challenges of the field. PMID:11673116

  1. Smart vision chips: An overview

    NASA Technical Reports Server (NTRS)

    Koch, Christof

    1994-01-01

    This viewgraph presentation presents four working analog VLSI vision chips: (1) time-derivative retina, (2) zero-crossing chip, (3) resistive fuse, and (4) figure-ground chip; work in progress on computing motion and neuromorphic systems; and conceptual and practical lessons learned.

  2. Chipping citrus wood for gasification

    SciTech Connect

    Churchill, D.B.; Hedden, S.L.; Whitney, J.D.; Shaw, L.N.

    1985-01-01

    Non-productive citrus trees were chipped with a portable fly-wheel-type chipper powered by a 45 kW engine. Chips were air dried under an open shed to 14% (w.b.) moisture content. By weight, approximately 50% of the total tree could be made into usable chips. The root system averaged 36% of the total tree weight.

  3. Functional independence of monomeric CHIP28 water channels revealed by expression of wild-type mutant heterodimers.

    PubMed

    Shi, L B; Skach, W R; Verkman, A S

    1994-04-01

    CHIP28 is a major water transporting protein in erythrocytes and kidney which forms tetramers in membranes (Verbavatz, J. M., Brown, D., Sabolic, I., Valenti, G., Ausiello, D. A., Van Hoek, A. N., Ma, T., and Verkman, A. S. (1993) J. Cell Biol. 123, 605-618). To determine whether CHIP28 monomers function independently, chimeric cDNA dimers were constructed which contained wild-type CHIP28 in series with either wild-type CHIP28, a non-water transporting CHIP28 mutant (C189W), or a functional but mercurial-insensitive CHIP28 mutant (C189S). Transcribed cRNAs were injected in Xenopus oocytes and plasma membrane expression was assayed by quantitative immunofluorescence. Water channel function was measured by osmotically induced swelling. CHIP28 homo- and heterodimers were targeted to the oocyte plasma membrane and functioned as water channels. Relative osmotic water permeability (Pf) values (normalized for plasma membrane expression of monomeric subunits) were: 1.0 (CHIP28 monomer), 0.0 (C189W), 1.07 (C189S), 1.10 (CHIP28-CHIP28 dimer) and 0.52 (CHIP28-C189W). The increase in oocyte Pf was linearly related to plasma membrane expression of wild-type CHIP28 and C189S subunits. HgCl2 (0.3 mM) inhibited channel-mediated Pf in oocytes expressing wild-type CHIP28 monomers and dimers by 85-90%, but did not inhibit Pf in oocytes expressing C189S. HgCl2 inhibited Pf in oocytes expressing CHIP28-C189S dimers by 44 +/- 7%, consistent with one mercurial-sensitive and one insensitive subunit in the heterodimer. These results indicate that despite their assembly in tetramers, monomeric CHIP28 subunits function independently as water channels. PMID:7511600

  4. World with Chips

    NASA Astrophysics Data System (ADS)

    Hoefflinger, Bernd

    Although we are well advised to look at the future 1 day at a time, we have seen in the chapters of this book, and they necessarily could cover only a selection on the features and applications of those tiny chips, that their potential continues to grow at the exceptional rates of the past. However, the new commitment has to be towards Sustainable Nanoelectronics, guided by creating sensing, computing, memory, and communication functions, which move just a few electrons per operation, each operation consuming energy less than one or a few femtojoule, less than any of the 1014 synapses in our brains. At these energy levels, chips can serve everywhere, making them ubiquitous, pervasive, certainly wireless, and often energy-autonomous. The expected six Billion users of these chips in 2020, through their mobile, intelligent companions, will benefit from global and largely equal access to information, education, knowledge, skills, and care.

  5. Cytometer on a Chip

    NASA Technical Reports Server (NTRS)

    Fernandez, Salvador M.

    2011-01-01

    A cytometer now under development exploits spatial sorting of sampled cells on a microarray chip followed by use of grating-coupled surface-plasmon-resonance imaging (GCSPRI) to detect the sorted cells. This cytometer on a chip is a prototype of contemplated future miniature cytometers that would be suitable for rapidly identifying pathogens and other cells of interest in both field and laboratory applications and that would be attractive as alternatives to conventional flow cytometers. The basic principle of operation of a conventional flow cytometer requires fluorescent labeling of sampled cells, stringent optical alignment of a laser beam with a narrow orifice, and flow of the cells through the orifice, which is subject to clogging. In contrast, the principle of operation of the present cytometer on a chip does not require fluorescent labeling of cells, stringent optical alignment, or flow through a narrow orifice. The basic principle of operation of the cytometer on a chip also reduces the complexity, mass, and power of the associated laser and detection systems, relative to those needed in conventional flow cytometry. Instead of making cells flow in single file through a narrow flow orifice for sequential interrogation as in conventional flow cytometry, a liquid containing suspended sampled cells is made to flow over the front surface of a microarray chip on which there are many capture spots. Each capture spot is coated with a thin (.50-nm) layer of gold that is, in turn, coated with antibodies that bind to cell-surface molecules characteristic of the cell species of interest. The multiplicity of capture spots makes it possible to perform rapid, massively parallel analysis of a large cell population. The binding of cells to each capture spot gives rise to a minute change in the index of refraction at the surface of the chip. This change in the index of refraction is what is sensed in GCSPRI, as described briefly below. The identities of the various species in

  6. Cytometer on a Chip

    NASA Technical Reports Server (NTRS)

    Fernandez, Salvador M.

    2011-01-01

    A cytometer now under development exploits spatial sorting of sampled cells on a microarray chip followed by use of grating-coupled surface-plasmon-resonance imaging (GCSPRI) to detect the sorted cells. This cytometer on a chip is a prototype of contemplated future miniature cytometers that would be suitable for rapidly identifying pathogens and other cells of interest in both field and laboratory applications and that would be attractive as alternatives to conventional flow cytometers. The basic principle of operation of a conventional flow cytometer requires fluorescent labeling of sampled cells, stringent optical alignment of a laser beam with a narrow orifice, and flow of the cells through the orifice, which is subject to clogging. In contrast, the principle of operation of the present cytometer on a chip does not require fluorescent labeling of cells, stringent optical alignment, or flow through a narrow orifice. The basic principle of operation of the cytometer on a chip also reduces the complexity, mass, and power of the associated laser and detection systems, relative to those needed in conventional flow cytometry. Instead of making cells flow in single file through a narrow flow orifice for sequential interrogation as in conventional flow cytometry, a liquid containing suspended sampled cells is made to flow over the front surface of a microarray chip on which there are many capture spots. Each capture spot is coated with a thin (approximately 50-nm) layer of gold that is, in turn, coated with antibodies that bind to cell-surface molecules characteristic of one the cell species of interest. The multiplicity of capture spots makes it possible to perform rapid, massively parallel analysis of a large cell population. The binding of cells to each capture spot gives rise to a minute change in the index of refraction at the surface of the chip. This change in the index of refraction is what is sensed in GCSPRI, as described briefly below. The identities of the

  7. Justification of rapid prototyping in the development cycle of thermoplastic-based lab-on-a-chip.

    PubMed

    Preywisch, Regina; Ritzi-Lehnert, Marion; Drese, Klaus S; Röser, Tina

    2011-11-01

    During the developmental cycle of lab-on-a-chip devices, various microstructuring techniques are required. While in the designing and assay implementation phase direct structuring or so-called rapid-prototyping methods such as milling or laser ablation are applied, replication methods like hot embossing or injection moulding are favourable for large quantity manufacturing. This work investigated the applicability of rapid-prototyping techniques for thermoplastic chip development in general, and the reproducibility of performances in dependency of the structuring technique. A previously published chip for prenatal diagnosis that preconcentrates DNA via electrokinetic trapping and field-amplified-sample-stacking and afterwards separates it in CGE was chosen as a model. The impact of structuring, sealing, and the integration of membranes on the mobility of the EOF, DNA preconcentration, and DNA separation was studied. Structuring methods were found to significantly change the location where preconcentration of DNA occurs. However, effects on the mobility of the EOF and the separation quality of DNA were not observed. Exchange of the membrane has no effect on the chip performance, whereas the sealing method impairs the separation of DNA within the chip. The overall assay performance is not significantly influenced by different structuring methods; thus, the application of rapid-prototyping methods during a chip development cycle is well justified. PMID:22102495

  8. DNA transformation via local heat shock

    NASA Astrophysics Data System (ADS)

    Li, Sha; Meadow Anderson, L.; Yang, Jui-Ming; Lin, Liwei; Yang, Haw

    2007-07-01

    This work describes transformation of foreign DNA into bacterial host cells by local heat shock using a microfluidic system with on-chip, built-in platinum heaters. Plasmid DNA encoding ampicillin resistance and a fluorescent protein can be effectively transformed into the DH5α chemically competent E. coli using this device. Results further demonstrate that only one-thousandth of volume is required to obtain transformation efficiencies as good as or better than conventional practices. As such, this work complements other lab-on-a-chip technologies for potential gene cloning/therapy and protein expression applications.

  9. Radiometer on a Chip

    NASA Technical Reports Server (NTRS)

    Chattopadhyay, Goutam; Gill, John J.; Mehdi, Imran; Lee, Choonsup; Schlecht, Erich T.; Skalare, Anders; Ward, John S.; Siegel, Peter H.; Thomas, Bertrand C.

    2009-01-01

    The radiometer on a chip (ROC) integrates whole wafers together to p rovide a robust, extremely powerful way of making submillimeter rece ivers that provide vertically integrated functionality. By integratin g at the wafer level, customizing the interconnects, and planarizing the transmission media, it is possible to create a lightweight asse mbly performing the function of several pieces in a more conventiona l radiometer.

  10. Genome-wide localization of Rrm3 and Pif1 DNA helicases at stalled active and inactive DNA replication forks of Saccharomyces cerevisiae.

    PubMed

    Rossi, Silvia Emma; Carotenuto, Walter; Giannattasio, Michele

    2016-03-01

    The genome of the budding yeast Saccharomyces cerevisiae is sequenced and the location and dynamic of activation of DNA replication origins are known. G1 synchronized yeast cells can be released into S-phase in the presence of hydroxyurea (HU) (1), which slows down DNA replication and retains replication forks in proximity of DNA replication origins. In this condition, the Chromatin Immuno-Precipitation on chip (ChIP on chip) (2-4) of replisome components allows the precise localization of all active DNA replication forks. This analysis can be coupled with the ssDNA-BromodeoxyUridine (ssDNA-BrdU) Immuno-Precipitation on chip (ssDNA-BrdU IP on chip) technique (5-7), which detects the location of newly synthesized DNA. Comparison of binding and BrdU incorporation profiles allows to locate a factor of interest at DNA replication forks genome wide. We present datasets deposited in the gene expression omnibus (GEO) database under accession number GSE68214, which show how the DNA helicases Rrm3 and Pif1 (8) associate to active and inactive DNA replication forks.

  11. High-throughput metabolic genotoxicity screening with a fluidic microwell chip and electrochemiluminescence†

    PubMed Central

    Wasalathanthri, Dhanuka P.; Malla, Spundana; Bist, Itti; Tang, Chi K.; Faria, Ronaldo C.; Rusling, James F.

    2014-01-01

    A high throughput electrochemiluminescent (ECL) chip was fabricated and integrated into a fluidic system for screening toxicity-related chemistry of drug and pollutant metabolites. The chip base is conductive pyrolytic graphite onto which are printed 64 microwells capable of holding one-µL droplets. Films combining DNA, metabolic enzymes and an ECL-generating ruthenium metallopolymer (RuIIPVP) are fabricated in these microwells. The system runs metabolic enzyme reactions, and subsequently detects DNA damage caused by reactive metabolites. The performance of the chip was tested by measuring DNA damage caused by metabolites of the well-known procarcinogen benzo[a]pyrene (B[a]P). Liver microsomes and cytochrome P450 (cyt P450) enzymes were used with and without epoxide hydrolase (EH), a conjugative enzyme required for multi-enzyme bioactivation of B[a]P. DNA adduct formation was confirmed by determining specific DNA-metabolite adducts using similar films of DNA/enzyme on magnetic bead biocolloid reactors, hydrolyzing the DNA, and analyzing by capillary liquid chromatography-mass spectrometry (CapLC-MS/MS). The fluidic chip was also used to measure IC50-values of inhibitors of cyt P450s. All results show good correlation with reported enzyme activity and inhibition assays. PMID:24113555

  12. Sodium chloride in supercritical water as a function of density: potentials of mean force and an equation for the dissociation constant from 723 to 1073 K and from 0 to 0.9 g/cm(3).

    PubMed

    Liu, Wenbin; Wood, Robert H; Doren, Douglas J

    2008-06-19

    The potential of mean force (PMF) of sodium chloride in water has been calculated by using the ab initio classical free-energy perturbation method at five state points: at 973 K with densities of 0.2796, 0.0935, and 0.0101 g/cm (3) and at 723 K with densities of 0.0897 and 0.0098 g/cm (3). The method is based on a QM-MM model in which Na-H 2O, Cl-H 2O, and Na-Cl interactions are calculated by ab initio methods. The water-water interactions are from the polarizable TIP4P-FQ model. The logarithm of the dissociation constant (log K c) has been calculated from the PMF. These predictions, together with experimental measurements, were used to derive an equation for log K c at densities from 0 to 0.9 g/cm (3) and temperatures from 723 to 1073 K, as well as from 600 to 1073 K for densities from 0.29 g/cm (3) to 0.9 g/cm (3). Extrapolation of the present equation below 723 K for densities less than 0.29 g/cm (3) does not fit the experimental results. This is attributed to long-range changes in the local dielectric constant due to the high compressibility. Comparisons with previous predictions and simulations are presented.

  13. Sodium chloride in supercritical water as a function of density: potentials of mean force and an equation for the dissociation constant from 723 to 1073 K and from 0 to 0.9 g/cm(3).

    PubMed

    Liu, Wenbin; Wood, Robert H; Doren, Douglas J

    2008-06-19

    The potential of mean force (PMF) of sodium chloride in water has been calculated by using the ab initio classical free-energy perturbation method at five state points: at 973 K with densities of 0.2796, 0.0935, and 0.0101 g/cm (3) and at 723 K with densities of 0.0897 and 0.0098 g/cm (3). The method is based on a QM-MM model in which Na-H 2O, Cl-H 2O, and Na-Cl interactions are calculated by ab initio methods. The water-water interactions are from the polarizable TIP4P-FQ model. The logarithm of the dissociation constant (log K c) has been calculated from the PMF. These predictions, together with experimental measurements, were used to derive an equation for log K c at densities from 0 to 0.9 g/cm (3) and temperatures from 723 to 1073 K, as well as from 600 to 1073 K for densities from 0.29 g/cm (3) to 0.9 g/cm (3). Extrapolation of the present equation below 723 K for densities less than 0.29 g/cm (3) does not fit the experimental results. This is attributed to long-range changes in the local dielectric constant due to the high compressibility. Comparisons with previous predictions and simulations are presented. PMID:18491938

  14. A chromatin immunoprecipitation (ChIP) protocol for use in whole human adipose tissue.

    PubMed

    Haim, Yulia; Tarnovscki, Tanya; Bashari, Dana; Rudich, Assaf

    2013-11-01

    Chromatin immunoprecipitation (ChIP) has become a central method when studying in vivo protein-DNA interactions, with the major challenge being the hope to capture "authentic" interactions. While ChIP protocols have been optimized for use with specific cell types and tissues including adipose tissue-derived cells, a working ChIP protocol addressing the challenges imposed by fresh whole human adipose tissue has not been described. Utilizing human paired omental and subcutaneous adipose tissue obtained during elective abdominal surgeries, we have carefully identified and optimized individual steps in the ChIP protocol employed directly on fresh tissue fragments. We describe a complete working protocol for using ChIP on whole adipose tissue fragments. Specific steps required adaptation of the ChIP protocol to human whole adipose tissue. In particular, a cross-linking step was performed directly on fresh small tissue fragments. Nuclei were isolated before releasing chromatin, allowing better management of fat content; a sonication protocol to obtain fragmented chromatin was optimized. We also demonstrate the high sensitivity of immunoprecipitated chromatin from adipose tissue to freezing. In conclusion, we describe the development of a ChIP protocol optimized for use in studying whole human adipose tissue, providing solutions for the unique challenges imposed by this tissue. Unraveling protein-DNA interaction in whole human adipose tissue will likely contribute to elucidating molecular pathways contributing to common human diseases such as obesity and type 2 diabetes.

  15. Microfluidic Devices for Forensic DNA Analysis: A Review.

    PubMed

    Bruijns, Brigitte; van Asten, Arian; Tiggelaar, Roald; Gardeniers, Han

    2016-01-01

    Microfluidic devices may offer various advantages for forensic DNA analysis, such as reduced risk of contamination, shorter analysis time and direct application at the crime scene. Microfluidic chip technology has already proven to be functional and effective within medical applications, such as for point-of-care use. In the forensic field, one may expect microfluidic technology to become particularly relevant for the analysis of biological traces containing human DNA. This would require a number of consecutive steps, including sample work up, DNA amplification and detection, as well as secure storage of the sample. This article provides an extensive overview of microfluidic devices for cell lysis, DNA extraction and purification, DNA amplification and detection and analysis techniques for DNA. Topics to be discussed are polymerase chain reaction (PCR) on-chip, digital PCR (dPCR), isothermal amplification on-chip, chip materials, integrated devices and commercially available techniques. A critical overview of the opportunities and challenges of the use of chips is discussed, and developments made in forensic DNA analysis over the past 10-20 years with microfluidic systems are described. Areas in which further research is needed are indicated in a future outlook.

  16. Microfluidic Devices for Forensic DNA Analysis: A Review

    PubMed Central

    Bruijns, Brigitte; van Asten, Arian; Tiggelaar, Roald; Gardeniers, Han

    2016-01-01

    Microfluidic devices may offer various advantages for forensic DNA analysis, such as reduced risk of contamination, shorter analysis time and direct application at the crime scene. Microfluidic chip technology has already proven to be functional and effective within medical applications, such as for point-of-care use. In the forensic field, one may expect microfluidic technology to become particularly relevant for the analysis of biological traces containing human DNA. This would require a number of consecutive steps, including sample work up, DNA amplification and detection, as well as secure storage of the sample. This article provides an extensive overview of microfluidic devices for cell lysis, DNA extraction and purification, DNA amplification and detection and analysis techniques for DNA. Topics to be discussed are polymerase chain reaction (PCR) on-chip, digital PCR (dPCR), isothermal amplification on-chip, chip materials, integrated devices and commercially available techniques. A critical overview of the opportunities and challenges of the use of chips is discussed, and developments made in forensic DNA analysis over the past 10–20 years with microfluidic systems are described. Areas in which further research is needed are indicated in a future outlook. PMID:27527231

  17. Microfluidic Devices for Forensic DNA Analysis: A Review.

    PubMed

    Bruijns, Brigitte; van Asten, Arian; Tiggelaar, Roald; Gardeniers, Han

    2016-01-01

    Microfluidic devices may offer various advantages for forensic DNA analysis, such as reduced risk of contamination, shorter analysis time and direct application at the crime scene. Microfluidic chip technology has already proven to be functional and effective within medical applications, such as for point-of-care use. In the forensic field, one may expect microfluidic technology to become particularly relevant for the analysis of biological traces containing human DNA. This would require a number of consecutive steps, including sample work up, DNA amplification and detection, as well as secure storage of the sample. This article provides an extensive overview of microfluidic devices for cell lysis, DNA extraction and purification, DNA amplification and detection and analysis techniques for DNA. Topics to be discussed are polymerase chain reaction (PCR) on-chip, digital PCR (dPCR), isothermal amplification on-chip, chip materials, integrated devices and commercially available techniques. A critical overview of the opportunities and challenges of the use of chips is discussed, and developments made in forensic DNA analysis over the past 10-20 years with microfluidic systems are described. Areas in which further research is needed are indicated in a future outlook. PMID:27527231

  18. Chip-Based Sensors for Disease Diagnosis

    NASA Astrophysics Data System (ADS)

    Fang, Zhichao

    Nucleic acid analysis is one of the most important disease diagnostic approaches in medical practice, and has been commonly used in cancer biomarker detection, bacterial speciation and many other fields in laboratory. Currently, the application of powerful research methods for genetic analysis, including the polymerase chain reaction (PCR), DNA sequencing, and gene expression profiling using fluorescence microarrays, are not widely used in hospitals and extended-care units due to high-cost, long detection times, and extensive sample preparation. Bioassays, especially chip-based electrochemical sensors, may be suitable for the next generation of rapid, sensitive, and multiplexed detection tools. Herein, we report three different microelectrode platforms with capabilities enabled by nano- and microtechnology: nanoelectrode ensembles (NEEs), nanostructured microelectrodes (NMEs), and hierarchical nanostructured microelectrodes (HNMEs), all of which are able to directly detect unpurified RNA in clinical samples without enzymatic amplification. Biomarkers that are cancer and infectious disease relevant to clinical medicine were chosen to be the targets. Markers were successfully detected with clinically-relevant sensitivity. Using peptide nucleic acids (PNAs) as probes and an electrocatalytic reporter system, NEEs were able to detect prostate cancer-related gene fusions in tumor tissue samples with 100 ng of RNA. The development of NMEs improved the sensitivity of the assay further to 10 aM of DNA target, and multiplexed detection of RNA sequences of different prostate cancer-related gene fusion types was achieved on the chip-based NMEs platform. An HNMEs chip integrated with a bacterial lysis device was able to detect as few as 25 cfu bacteria in 30 minutes and monitor the detection in real time. Bacterial detection could also be performed in neat urine samples. The development of these versatile clinical diagnostic tools could be extended to the detection of various

  19. Circulating polymerase chain reaction chips utilizing multiple-membrane activation

    NASA Astrophysics Data System (ADS)

    Wang, Chih-Hao; Chen, Yi-Yu; Liao, Chia-Sheng; Hsieh, Tsung-Min; Luo, Ching-Hsing; Wu, Jiunn-Jong; Lee, Huei-Huang; Lee, Gwo-Bin

    2007-02-01

    This paper reports a new micromachined, circulating, polymerase chain reaction (PCR) chip for nucleic acid amplification. The PCR chip is comprised of a microthermal control module and a polydimethylsiloxane (PDMS)-based microfluidic control module. The microthermal control modules are formed with three individual heating and temperature-sensing sections, each modulating a specific set temperature for denaturation, annealing and extension processes, respectively. Micro-pneumatic valves and multiple-membrane activations are used to form the microfluidic control module to transport sample fluids through three reaction regions. Compared with other PCR chips, the new chip is more compact in size, requires less time for heating and cooling processes, and has the capability to randomly adjust time ratios and cycle numbers depending on the PCR process. Experimental results showed that detection genes for two pathogens, Streptococcus pyogenes (S. pyogenes, 777 bps) and Streptococcus pneumoniae (S. pneumoniae, 273 bps), can be successfully amplified using the new circulating PCR chip. The minimum number of thermal cycles to amplify the DNA-based S. pyogenes for slab gel electrophoresis is 20 cycles with an initial concentration of 42.5 pg µl-1. Experimental data also revealed that a high reproducibility up to 98% could be achieved if the initial template concentration of the S. pyogenes was higher than 4 pg µl-1. The preliminary results of the current paper were presented at the 19th IEEE International Conference on Micro Electro Mechanical Systems (IEEE MEMS 2006), Istanbul, Turkey, 22-26 January, 2006.

  20. Phospholipid Polymer Biointerfaces for Lab-on-a-Chip Devices.

    PubMed

    Xu, Yan; Takai, Madoka; Ishihara, Kazuhiko

    2010-06-01

    This review summarizes recent achievements and progress in the development of various functional 2-methacryloyloxyethyl phosphorylcholine (MPC) polymer biointerfaces for lab-on-a-chip devices and applications. As phospholipid polymers, MPC polymers can form cell-membrane-like surfaces by surface chemistry and physics and thereby provide biointerfaces capable of suppressing protein adsorption and many subsequent biological responses. In order to enable application to microfluidic devices, a number of MPC polymers with diverse functions have been specially designed and synthesized by incorporating functional units such as charge and active ester for generating the microfluidic flow and conjugating biomolecules, respectively. Furthermore, these polymers were incorporated with silane or hydrophobic moiety to construct stable interfaces on various substrate materials such as glass, quartz, poly(methyl methacrylate), and poly(dimethylsiloxane), via a silane-coupling reaction or hydrophobic interactions. The basic interfacial properties of these interfaces have been characterized from multiple aspects of chemistry, physics, and biology, and the suppression of nonspecific bioadsorption and control of microfluidic flow have been successfully achieved using these biointerfaces on a chip. Further, many chip-based biomedical applications such as immunoassays and DNA separation have been accomplished by integrating these biointerfaces on a chip. Therefore, functional phospholipid polymer interfaces are promising and useful for application to lab-on-a-chip devices in biomedicine.

  1. Acoustic micro-vortexing of fluids, particles and cells in disposable microfluidic chips.

    PubMed

    Iranmanesh, Ida; Ohlin, Mathias; Ramachandraiah, Harisha; Ye, Simon; Russom, Aman; Wiklund, Martin

    2016-08-01

    We demonstrate an acoustic platform for micro-vortexing in disposable polymer microfluidic chips with small-volume (20 μl) reaction chambers. The described method is demonstrated for a variety of standard vortexing functions, including mixing of fluids, re-suspension of a pellet of magnetic beads collected by a magnet placed on the chip, and lysis of cells for DNA extraction. The device is based on a modified Langevin-type ultrasonic transducer with an exponential horn for efficient coupling into the microfluidic chip, which is actuated by a low-cost fixed-frequency electronic driver board. The transducer is optimized by numerical modelling, and different demonstrated vortexing functions are realized by actuating the transducer for varying times; from fractions of a second for fluid mixing, to half a minute for cell lysis and DNA extraction. The platform can be operated during 1 min below physiological temperatures with the help of a PC fan, a Peltier element and an aluminum heat sink acting as the chip holder. As a proof of principle for sample preparation applications, we demonstrate on-chip cell lysis and DNA extraction within 25 s. The method is of interest for automating and chip-integrating sample preparation procedures in various biological assays. PMID:27444649

  2. Acoustic micro-vortexing of fluids, particles and cells in disposable microfluidic chips.

    PubMed

    Iranmanesh, Ida; Ohlin, Mathias; Ramachandraiah, Harisha; Ye, Simon; Russom, Aman; Wiklund, Martin

    2016-08-01

    We demonstrate an acoustic platform for micro-vortexing in disposable polymer microfluidic chips with small-volume (20 μl) reaction chambers. The described method is demonstrated for a variety of standard vortexing functions, including mixing of fluids, re-suspension of a pellet of magnetic beads collected by a magnet placed on the chip, and lysis of cells for DNA extraction. The device is based on a modified Langevin-type ultrasonic transducer with an exponential horn for efficient coupling into the microfluidic chip, which is actuated by a low-cost fixed-frequency electronic driver board. The transducer is optimized by numerical modelling, and different demonstrated vortexing functions are realized by actuating the transducer for varying times; from fractions of a second for fluid mixing, to half a minute for cell lysis and DNA extraction. The platform can be operated during 1 min below physiological temperatures with the help of a PC fan, a Peltier element and an aluminum heat sink acting as the chip holder. As a proof of principle for sample preparation applications, we demonstrate on-chip cell lysis and DNA extraction within 25 s. The method is of interest for automating and chip-integrating sample preparation procedures in various biological assays.

  3. On-Chip Biomedical Imaging

    PubMed Central

    Göröcs, Zoltán; Ozcan, Aydogan

    2012-01-01

    Lab-on-a-chip systems have been rapidly emerging to pave the way toward ultra-compact, efficient, mass producible and cost-effective biomedical research and diagnostic tools. Although such microfluidic and micro electromechanical systems achieved high levels of integration, and are capable of performing various important tasks on the same chip, such as cell culturing, sorting and staining, they still rely on conventional microscopes for their imaging needs. Recently several alternative on-chip optical imaging techniques have been introduced, which have the potential to substitute conventional microscopes for various lab-on-a-chip applications. Here we present a critical review of these recently emerging on-chip biomedical imaging modalities, including contact shadow imaging, lensfree holographic microscopy, fluorescent on-chip microscopy and lensfree optical tomography. PMID:23558399

  4. Forensic Analysis of BIOS Chips

    NASA Astrophysics Data System (ADS)

    Gershteyn, Pavel; Davis, Mark; Shenoi, Sujeet

    Data can be hidden in BIOS chips without hindering computer performance. This feature has been exploited by virus writers and computer game enthusiasts. Unused BIOS storage can also be used by criminals, terrorists and intelligence agents to conceal secrets. However, BIOS chips are largely ignored in digital forensic investigations. Few techniques exist for imaging BIOS chips and no tools are available specifically for analyzing BIOS data.

  5. Securing Contactless Chips with PACE

    NASA Astrophysics Data System (ADS)

    Kügler, Dennis

    PACE (Password Authenticated Connection Establishment) is a cryptographic protocol that was developed to provide a secure knowledge-based authentication mechanism for contactless chips. The problems that are inherent to (but not limited to) contactless chips are described and PACE as a solution based on cryptographic tools is sketched. Finally, it is shown how to use PACE together with traditional short PINs of 4-6 digits as access control mechanism for contactless chips withstanding denial-of-service attacks.

  6. Nanoparticle Reactions on Chip

    NASA Astrophysics Data System (ADS)

    Köhler, J. M.; Kirner, Th.; Wagner, J.; Csáki, A.; Möller, R.; Fritzsche, W.

    The handling of heterogenous systems in micro reactors is difficult due to their adhesion and transport behaviour. Therefore, the formation of precipitates and gas bubbles has to be avoided in micro reaction technology, in most cases. But, micro channels and other micro reactors offer interesting possibilities for the control of reaction conditions and transport by diffusion and convection due to the laminar flow caused by small Reynolds numbers. This can be used for the preparation and modification of objects, which are much smaller than the cross section of microchannels. The formation of colloidal solutions and the change of surface states of nano particles are two important tasks for the application of chip reactors in nanoparticle technology. Some concepts for the preparation and reaction of nanoparticles in modular chip reactor arrangements will be discussed.

  7. The Human MitoChip: a high-throughput sequencing microarray for mitochondrial mutation detection.

    PubMed

    Maitra, Anirban; Cohen, Yoram; Gillespie, Susannah E D; Mambo, Elizabeth; Fukushima, Noriyoshi; Hoque, Mohammad O; Shah, Nila; Goggins, Michael; Califano, Joseph; Sidransky, David; Chakravarti, Aravinda

    2004-05-01

    Somatic mitochondrial mutations are common in human cancers, and can be used as a tool for early detection of cancer. We have developed a mitochondrial Custom Reseq microarray as an array-based sequencing platform for rapid and high-throughput analysis of mitochondrial DNA. The MitoChip contains oligonucleotide probes synthesized using standard photolithography and solid-phase synthesis, and is able to sequence >29 kb of double-stranded DNA in a single assay. Both strands of the entire human mitochondrial coding sequence (15,451 bp) are arrayed on the MitoChip; both strands of an additional 12,935 bp (84% of coding DNA) are arrayed in duplicate. We used 300 ng of genomic DNA to amplify the mitochondrial coding sequence in three overlapping long PCR fragments. We then sequenced >2 million base pairs of mitochondrial DNA, and successfully assigned base calls at 96.0% of nucleotide positions. Replicate experiments demonstrated >99.99% reproducibility. In matched fluid samples (urine and pancreatic juice, respectively) obtained from five patients with bladder cancer and four with pancreatic cancer, the MitoChip detected at least one cancer-associated mitochondrial mutation in six (66%) of nine samples. The MitoChip is a high-throughput sequencing tool for the reliable identification of mitochondrial DNA mutations from primary tumors in clinical samples.

  8. The detection of p53 gene via fluorescence quenching of quantum dot in microfluidic chip.

    PubMed

    Yoo, Jeong Ha; Yoo, In Sang; Yoon, Won Jung; Kim, Jong Sung

    2012-05-01

    Recently, quantum dot (QD) has been used widely in the field of bio assay including cell imaging, biomarker, and fluorescence resonance energy transfer (FRET) sensor. The DNA assay without labeling process has several advantages including low cost, short time, and simplicity. Microbeads of agarose, glass, and polystyrene have been used as a solid support in microfluidic devices to trace molecules. The main advantages of microfluidics include high throughput, short analysis time, small sample volume, and high sensitivity. PDMS based microfluidic chips were prepared for the detection of p53 gene by using QD-DNA conjugate. The microfluidic chip has a weir in the channel to trap microbeads to which QD-DNA probes bind. Carboxylated CdSe/ZnS QDs (wavelength of emission: 605 nm) could bind to microbeads of polystyrene/divinyl benzene via EDC/NHS crosslinking reaction. The target gene and DNA intercalating dye (TOTO-3) were loaded into the micro-channel. Fluorescence quenching from QDs by intercalating dye was observed after hybridization of DNA at the weir in the channel of microfluidic chip. The fluorescence quenching from QDs by TOTO-3 was dependent on the concentration of target gene. This experiment shows the possibility of rapid detection of DNA via bead-QD complex on microfluidic chip. PMID:22852354

  9. Lab-on-a-chip PCR: real time PCR in miniaturized format for HLA diagnostics

    NASA Astrophysics Data System (ADS)

    Gaertner, Claudia; Becker, Holger; Hlawatsch, Nadine; Klemm, Richard; Moche, Christian; Sewart, René; Frank, Rainer; Willems, Andreas

    2014-05-01

    In case of transplantation or the identification of special metabolic diseases like coeliac disease, HLA typing has to be done fast and reliably with easy-to-handle devices by using limited amount of sample. Against this background a lab-on-a-chip device was realized enabling a fast HLA typing via miniaturized Real-time PCR. Hereby, two main process steps were combined, namely the extraction of DNA from whole blood and the amplification of the target DNA by Real-time PCR giving rise-to a semi-quantitative analysis. For the implementation of both processes on chip, a sample preparation and a real-time module were used. Sample preparation was carried out by using magnetic beads that were stored directly on chip as dry powder, together with all lysis reagents. After purification of the DNA by applying a special buffer regime, the sample DNA was transferred into the PCR module for amplification and detection. Coping with a massively increased surface-to-volume ratio, which results in a higher amount of unspecific binding on the chip surface, special additives needed to be integrated to compensate for this effect. Finally the overall procedure showed a sensitivity comparable to standard Real-time PCR but reduced the duration of analysis to significantly less than one hour. The presented work demonstrates that the combination of lab-on-a-chip PCR with direct optical read-out in a real-time fashion is an extremely promising tool for molecular diagnostics.

  10. SR proteins Asf/SF2 and 9G8 interact to activate enhancer-dependent intron D splicing of bovine growth hormone pre-mRNA in vitro.

    PubMed Central

    Li, X; Shambaugh, M E; Rottman, F M; Bokar, J A

    2000-01-01

    The alternative splicing of the last intron (intron D) of bovine growth hormone (bGH) pre-mRNA requires a down-stream exonic splicing enhancer (FP/ESE). The presence of at least one SR protein has been shown to be essential for FP/ESE function and splicing of intron D in in vitro splicing assays. However, in vitro reconstitution of splicing using individual purified SR proteins may not accurately reflect the true complexity of alternative splicing in an intact nucleus, where multiple SR proteins in varying amounts are likely to be available simultaneously. Here, a panel of recombinant baculovirus-expressed SR proteins was produced and tested for the ability to activate FP/ESE-dependent splicing. Individual recombinant SR proteins differed significantly in their activity in promoting intron D splicing. Among the recombinant SR proteins tested, SRp55 was the most active, SC35 showed very little activity, and ASF/SF2 and 9G8 individually had intermediate activity. At least one SR protein (ASF/SF2) bound to the FP/ESE with characteristics of a cooperative interaction. Most interestingly, low concentrations of ASF/SF2 and 9G8 acted synergistically to activate intron D splicing. This was due in part to synergistic binding to the FP/ESE. Splicing of bGH intron D is inherently complex, and is likely controlled by an interaction of the FP/ESE with several trans-acting protein factors acting both independently and cooperatively. This level of complexity may be required for precise control of alternative splicing by an exon sequence, which simultaneously is constrained to maintain translational integrity of the mature mRNA. PMID:11142383

  11. Cell lysis and DNA extraction in microfabricated devices

    NASA Astrophysics Data System (ADS)

    Prinz, Christelle; Tegenfeldt, Jonas; Austin, Robert

    2002-03-01

    We are developing a microfabricated device to lyse single cells and extract the DNA. The chip consists of two parts: a diffuse mixer combined with a dielectrophoretic trap. We are working with E. coli which have been made osmoticaly unstable before loading into the chip. The cells are lysed by osmotic shock in the mixer. The lysate is then passed to the dielectrophoretic trap. Attempts to separate the genomic DNA from the lysate fragments by selectively trapping the DNA using dielectrophoresis have been made. We have encountered cell sticking problems and are investingating surface modifications using Polyethylene glycol to solve this problem.

  12. Plasmonic SERS biosensing nanochips for DNA detection.

    PubMed

    Ngo, Hoan T; Wang, Hsin-Neng; Fales, Andrew M; Vo-Dinh, Tuan

    2016-03-01

    The development of rapid, cost-effective DNA detection methods for molecular diagnostics at the point-of-care (POC) has been receiving increasing interest. This article reviews several DNA detection techniques based on plasmonic-active nanochip platforms developed in our laboratory over the last 5 years, including the molecular sentinel-on-chip (MSC), the multiplex MSC, and the inverse molecular sentinel-on-chip (iMS-on-Chip). DNA probes were used as the recognition elements, and surface-enhanced Raman scattering (SERS) was used as the signal detection method. Sensing mechanisms were based on hybridization of target sequences and DNA probes, resulting in a distance change between SERS reporters and the nanochip's plasmonic-active surface. As the field intensity of the surface plasmon decays exponentially as a function of distance, the distance change in turn affects SERS signal intensity, thus indicating the presence and capture of the target sequences. Our techniques were single-step DNA detection techniques. Target sequences were detected by simple delivery of sample solutions onto DNA probe-functionalized nanochips and measuring the SERS signal after appropriate incubation times. Target sequence labeling or washing to remove unreacted components was not required, making the techniques simple, easy-to-use, and cost-effective. The usefulness of the nanochip platform-based techniques for medical diagnostics was illustrated by the detection of host genetic biomarkers for respiratory viral infection and of the dengue virus gene.

  13. Plasmonic SERS biosensing nanochips for DNA detection.

    PubMed

    Ngo, Hoan T; Wang, Hsin-Neng; Fales, Andrew M; Vo-Dinh, Tuan

    2016-03-01

    The development of rapid, cost-effective DNA detection methods for molecular diagnostics at the point-of-care (POC) has been receiving increasing interest. This article reviews several DNA detection techniques based on plasmonic-active nanochip platforms developed in our laboratory over the last 5 years, including the molecular sentinel-on-chip (MSC), the multiplex MSC, and the inverse molecular sentinel-on-chip (iMS-on-Chip). DNA probes were used as the recognition elements, and surface-enhanced Raman scattering (SERS) was used as the signal detection method. Sensing mechanisms were based on hybridization of target sequences and DNA probes, resulting in a distance change between SERS reporters and the nanochip's plasmonic-active surface. As the field intensity of the surface plasmon decays exponentially as a function of distance, the distance change in turn affects SERS signal intensity, thus indicating the presence and capture of the target sequences. Our techniques were single-step DNA detection techniques. Target sequences were detected by simple delivery of sample solutions onto DNA probe-functionalized nanochips and measuring the SERS signal after appropriate incubation times. Target sequence labeling or washing to remove unreacted components was not required, making the techniques simple, easy-to-use, and cost-effective. The usefulness of the nanochip platform-based techniques for medical diagnostics was illustrated by the detection of host genetic biomarkers for respiratory viral infection and of the dengue virus gene. PMID:26547189

  14. Single chip camera active pixel sensor

    NASA Technical Reports Server (NTRS)

    Shaw, Timothy (Inventor); Pain, Bedabrata (Inventor); Olson, Brita (Inventor); Nixon, Robert H. (Inventor); Fossum, Eric R. (Inventor); Panicacci, Roger A. (Inventor); Mansoorian, Barmak (Inventor)

    2003-01-01

    A totally digital single chip camera includes communications to operate most of its structure in serial communication mode. The digital single chip camera include a D/A converter for converting an input digital word into an analog reference signal. The chip includes all of the necessary circuitry for operating the chip using a single pin.

  15. A nanoliter self-priming compartmentalization chip for point-of-care digital PCR analysis.

    PubMed

    Song, Qi; Gao, Yibo; Zhu, Qiangyuan; Tian, Qingchang; Yu, Bingwen; Song, Bofan; Xu, Yanan; Yuan, Maokai; Ma, Congcong; Jin, Wei; Zhang, Tao; Mu, Ying; Jin, Qinhan

    2015-01-01

    A nanoliter self-priming compartmentalization (SPC) microfluidic chip suited for the digital polymerase chain reaction (dPCR) analysis in point-of-care testing (POCT) has been developed. This dPCR chip is fabricated of polydimethylsiloxane (PDMS). After the dPCR chip is evacuated, there will be a negative pressure environment in the chip because of the gas solubility of PDMS. The negative pressure environment can provide a self-priming power so that the sample solutions can be sucked into each reaction chamber sequentially. The whole sampling process requires no external power and is valve-free. Channels that contain water are designed around each sample panel to prevent the solvent (water) from evaporating during dPCR process. A glass coverslip is also used as a waterproof layer, which is more convenient and more efficient than other waterproof methods seen in literature. This dPCR chip allows three samples to be amplified at the same time. Each sample is distributed into 1040 reaction chambers, and each chamber is only 2.08 nL. Human β-actin DNA solutions of known concentrations are used as the templates for the dPCR analyses to verify the sensitivity and accuracy of the method. Template DNA solutions diluted to concentrations of 300, 100 and 10 copies/μL are tested and shown that this simple, portable and self-priming dPCR chip can be used at any clinic as a real POCT technique. PMID:26022215

  16. A nanoliter self-priming compartmentalization chip for point-of-care digital PCR analysis.

    PubMed

    Song, Qi; Gao, Yibo; Zhu, Qiangyuan; Tian, Qingchang; Yu, Bingwen; Song, Bofan; Xu, Yanan; Yuan, Maokai; Ma, Congcong; Jin, Wei; Zhang, Tao; Mu, Ying; Jin, Qinhan

    2015-01-01

    A nanoliter self-priming compartmentalization (SPC) microfluidic chip suited for the digital polymerase chain reaction (dPCR) analysis in point-of-care testing (POCT) has been developed. This dPCR chip is fabricated of polydimethylsiloxane (PDMS). After the dPCR chip is evacuated, there will be a negative pressure environment in the chip because of the gas solubility of PDMS. The negative pressure environment can provide a self-priming power so that the sample solutions can be sucked into each reaction chamber sequentially. The whole sampling process requires no external power and is valve-free. Channels that contain water are designed around each sample panel to prevent the solvent (water) from evaporating during dPCR process. A glass coverslip is also used as a waterproof layer, which is more convenient and more efficient than other waterproof methods seen in literature. This dPCR chip allows three samples to be amplified at the same time. Each sample is distributed into 1040 reaction chambers, and each chamber is only 2.08 nL. Human β-actin DNA solutions of known concentrations are used as the templates for the dPCR analyses to verify the sensitivity and accuracy of the method. Template DNA solutions diluted to concentrations of 300, 100 and 10 copies/μL are tested and shown that this simple, portable and self-priming dPCR chip can be used at any clinic as a real POCT technique. PMID:26029750

  17. Repairable chip bonding/interconnect process

    DOEpatents

    Bernhardt, Anthony F.; Contolini, Robert J.; Malba, Vincent; Riddle, Robert A.

    1997-01-01

    A repairable, chip-to-board interconnect process which addresses cost and testability issues in the multi-chip modules. This process can be carried out using a chip-on-sacrificial-substrate technique, involving laser processing. This process avoids the curing/solvent evolution problems encountered in prior approaches, as well is resolving prior plating problems and the requirements for fillets. For repairable high speed chip-to-board connection, transmission lines can be formed on the sides of the chip from chip bond pads, ending in a gull wing at the bottom of the chip for subsequent solder.

  18. Repairable chip bonding/interconnect process

    DOEpatents

    Bernhardt, A.F.; Contolini, R.J.; Malba, V.; Riddle, R.A.

    1997-08-05

    A repairable, chip-to-board interconnect process which addresses cost and testability issues in the multi-chip modules is disclosed. This process can be carried out using a chip-on-sacrificial-substrate technique, involving laser processing. This process avoids the curing/solvent evolution problems encountered in prior approaches, as well is resolving prior plating problems and the requirements for fillets. For repairable high speed chip-to-board connection, transmission lines can be formed on the sides of the chip from chip bond pads, ending in a gull wing at the bottom of the chip for subsequent solder. 10 figs.

  19. Optimization of direct whole blood PCR amplification with applications on a static thermostat chip.

    PubMed

    Qu, Bai-Yan; Wu, Zhi-Yong; Tian, Xiao-Xi; Chen, Kun; Fang, Fang

    2007-11-01

    In this paper, direct whole blood PCR amplifications on a static chip thermostat without sample purifications are demonstrated; in these amplifications, problems such as cross-interferences and contaminations could be avoided. The amplification conditions, such as the compositions of reagents and thermal programs, were investigated systematically by a GeneAmp PCR system with a native p53 gene segment (about 543 bp) of human genome and an exterior lambda DNA segment (about 500 bp) as targets. Direct amplifications of p53 and K-ras (about 157 bp) gene segments from 0.5 microL blood samples were successfully demonstrated by a static PCR chip with an indium tin oxide glass substrate. The chip thermostat has a typical size of 25 mm x 25 mm, and a polyethylene tube was used as the PCR vial on the glass surface of the chip. Fuzzy proportional integration-differentiation algorithms were adopted in temperature controls of the chip with an aid of a micro-Pt100 sensor. In the direct PCR with the thermostat chip, the whole process only involves automatic thermal programs. This work demonstrated that a chip PCR for field test without desktop facilities is possible either for a point of care test or for forensic analysis.

  20. A Cytomorphic Chip for Quantitative Modeling of Fundamental Bio-Molecular Circuits.

    PubMed

    2015-08-01

    We describe a 0.35 μm BiCMOS silicon chip that quantitatively models fundamental molecular circuits via efficient log-domain cytomorphic transistor equivalents. These circuits include those for biochemical binding with automatic representation of non-modular and loading behavior, e.g., in cascade and fan-out topologies; for representing variable Hill-coefficient operation and cooperative binding; for representing inducer, transcription-factor, and DNA binding; for probabilistic gene transcription with analogic representations of log-linear and saturating operation; for gain, degradation, and dynamics of mRNA and protein variables in transcription and translation; and, for faithfully representing biological noise via tunable stochastic transistor circuits. The use of on-chip DACs and ADCs enables multiple chips to interact via incoming and outgoing molecular digital data packets and thus create scalable biochemical reaction networks. The use of off-chip digital processors and on-chip digital memory enables programmable connectivity and parameter storage. We show that published static and dynamic MATLAB models of synthetic biological circuits including repressilators, feed-forward loops, and feedback oscillators are in excellent quantitative agreement with those from transistor circuits on the chip. Computationally intensive stochastic Gillespie simulations of molecular production are also rapidly reproduced by the chip and can be reliably tuned over the range of signal-to-noise ratios observed in biological cells.

  1. chipD: a web tool to design oligonucleotide probes for high-density tiling arrays

    PubMed Central

    Dufour, Yann S.; Wesenberg, Gary E.; Tritt, Andrew J.; Glasner, Jeremy D.; Perna, Nicole T.; Mitchell, Julie C.; Donohue, Timothy J.

    2010-01-01

    chipD is a web server that facilitates design of DNA oligonucleotide probes for high-density tiling arrays, which can be used in a number of genomic applications such as ChIP-chip or gene-expression profiling. The server implements a probe selection algorithm that takes as an input, in addition to the target sequences, a set of parameters that allow probe design to be tailored to specific applications, protocols or the array manufacturer’s requirements. The algorithm optimizes probes to meet three objectives: (i) probes should be specific; (ii) probes should have similar thermodynamic properties; and (iii) the target sequence coverage should be homogeneous and avoid significant gaps. The output provides in a text format, the list of probe sequences with their genomic locations, targeted strands and hybridization characteristics. chipD has been used successfully to design tiling arrays for bacteria and yeast. chipD is available at http://chipd.uwbacter.org/. PMID:20529880

  2. DNA FROM ANCIENT STONE TOOLS AND BONES EXCAVATED AT BUGAS-HOLDING, WYOMING

    EPA Science Inventory

    Traces of DNA may preserve on ancient stone tools. We examined 24 chipped stone artifacts recovered from the Bugas-Holding site in northwestern Wyoming for the presence of DNA residues, and we compared DNA preservation in bones and stone tools from the same stratigraphic context...

  3. Teaching microfluidic diagnostics using Jell-O(®) chips.

    PubMed

    Yang, Cheng Wei T; Lagally, Eric T

    2013-01-01

    Microfluidics has emerged as a versatile technology that has found many applications, including DNA chips, fuel cells, and diagnostics. As the field of microfluidic diagnostics grows, it is important to introduce the principles of this technology to young students and the general public. The objective of this project was to create a simple and effective method that could be used to teach key microfluidics concepts using easily accessible materials. Similar to the poly(dimethylsiloxane) soft lithography technique, a Jell-O(®) "chip" is produced by pouring a mixture of Jell-O(®) and gelatine solution into a mold, which is constructed using foam plate, coffee stirrers, and double-sided tape. The plate is transferred to a 4°C refrigerator for curing, and then the Jell-O(®) chip is peeled off for experimental demonstrations. Three types of chips have been fabricated with different molds: a JELLO mold, a Y-channel mold, and a pH-sensor mold. Using these devices, the basics of microfluidic diagnostics can be demonstrated in one or two class periods. The method described in this chapter provides teachers with a fast and inexpensive way to introduce this technology, and students with a fun and hands-on way to understand the basics of microfluidic diagnostics. PMID:23329433

  4. Digital PCR on an integrated self-priming compartmentalization chip.

    PubMed

    Zhu, Qiangyuan; Qiu, Lin; Yu, Bingwen; Xu, Yanan; Gao, Yibo; Pan, Tingting; Tian, Qingchang; Song, Qi; Jin, Wei; Jin, Qinhan; Mu, Ying

    2014-03-21

    An integrated on-chip valve-free and power-free microfluidic digital PCR device is for the first time developed by making use of a novel self-priming compartmentalization and simple dehydration control to realize 'divide and conquer' for single DNA molecule detection. The high gas solubility of PDMS is exploited to provide the built-in power of self-priming so that the sample and oil are sequentially sucked into the device to realize sample self-compartmentalization based on surface tension. The lifespan of its self-priming capability was about two weeks tested using an air-tight packaging bottle sealed with a small amount of petroleum jelly, which is significant for a practical platform. The SPC chip contains 5120 independent 5 nL microchambers, allowing the samples to be compartmentalized completely. Using this platform, three different abundances of lung cancer related genes are detected to demonstrate the feasibility and flexibility of the microchip for amplifying a single nucleic acid molecule. For maximal accuracy, within less than 5% of the measurement deviation, the optimal number of positive chambers is between 400 and 1250 evaluated by the Poisson distribution, which means one panel can detect an average of 480 to 4804 template molecules. This device without world-to-chip connections eliminates the constraint of the complex pipeline control, and is an integrated on-chip platform, which would be a significant improvement to digital PCR automation and more user-friendly.

  5. Microfabricated polymer chip for capillary gel electrophoresis.

    PubMed

    Hong, J W; Hosokawa, K; Fujii, T; Seki, M; Endo, I

    2001-01-01

    A polymer (PDMS: poly(dimethylsiloxane)) microchip for capillary gel electrophoresis that can separate different sizes of DNA molecules in a small experimental scale is presented. This microchip can be easily produced by a simple PDMS molding method against a microfabricated master without the use of elaborate bonding processes. This PDMS microchip could be used as a single use device unlike conventional microchips made of glass, quartz or silicon. The capillary channel on the chip was partially filled with agarose gel that can enhance separation resolution of different sizes of DNA molecules and can shorten the channel length required for the separation of the sample compared to capillary electrophoresis in free-flow or polymer solution format. We discuss the optimal conditions for the gel preparation that could be used in the microchannel. DNA molecules were successfully driven by an electric field and separated to form bands in the range of 100 bp to 1 kbp in a 2.0% agarose-filled microchannel with 8 mm of effective separation length.

  6. A fully sealed plastic chip for multiplex PCR and its application in bacteria identification.

    PubMed

    Xu, Youchun; Yan, He; Zhang, Yan; Jiang, Kewei; Lu, Ying; Ren, Yonghong; Wang, Hui; Wang, Shan; Xing, Wanli

    2015-07-01

    Multiplex PCR is an effective tool for simultaneous multiple target detection but is limited by the intrinsic interference and competition among primer pairs when it is performed in one reaction tube. Dividing a multiplex PCR into many single PCRs is a simple strategy to overcome this issue. Here, we constructed a plastic, easy-to-use, fully sealed multiplex PCR chip based on reversible centrifugation for the simultaneous detection of 63 target DNA sequences. The structure of the chip is quite simple, which contains sine-shaped infusing channels and a number of reaction chambers connecting to one side of these channels. Primer pairs for multiplex PCR were sequentially preloaded in the different reaction chambers, and the chip was enclosed with PCR-compatible adhesive tape. For usage, the PCR master mix containing a DNA template is pipetted into the infusing channels and centrifuged into the reaction chambers, leaving the infusing channels filled with air to avoid cross-contamination of the different chambers. Then, the chip is sealed and placed on a flat thermal cycler for PCR. Finally, amplification products can be detected in situ using a fluorescence scanner or recovered by reverse centrifugation for further analyses. Therefore, our chip possesses two functions: 1) it can be used for multi-target detection based on end-point in situ fluorescence detection; and 2) it can work as a sample preparation unit for analyses that need multiplex PCR such as hybridization and target sequencing. The performance of this chip was carefully examined and further illustrated in the identification of 8 pathogenic bacterial genomic DNA samples and 13 drug-resistance genes. Due to simplicity of its structure and operation, accuracy and generality, high-throughput capacity, and versatile functions (i.e., for in situ detection and sample preparation), our multiplex PCR chip has great potential in clinical diagnostics and nucleic acid-based point-of-care testing. PMID:26016439

  7. A fully sealed plastic chip for multiplex PCR and its application in bacteria identification.

    PubMed

    Xu, Youchun; Yan, He; Zhang, Yan; Jiang, Kewei; Lu, Ying; Ren, Yonghong; Wang, Hui; Wang, Shan; Xing, Wanli

    2015-07-01

    Multiplex PCR is an effective tool for simultaneous multiple target detection but is limited by the intrinsic interference and competition among primer pairs when it is performed in one reaction tube. Dividing a multiplex PCR into many single PCRs is a simple strategy to overcome this issue. Here, we constructed a plastic, easy-to-use, fully sealed multiplex PCR chip based on reversible centrifugation for the simultaneous detection of 63 target DNA sequences. The structure of the chip is quite simple, which contains sine-shaped infusing channels and a number of reaction chambers connecting to one side of these channels. Primer pairs for multiplex PCR were sequentially preloaded in the different reaction chambers, and the chip was enclosed with PCR-compatible adhesive tape. For usage, the PCR master mix containing a DNA template is pipetted into the infusing channels and centrifuged into the reaction chambers, leaving the infusing channels filled with air to avoid cross-contamination of the different chambers. Then, the chip is sealed and placed on a flat thermal cycler for PCR. Finally, amplification products can be detected in situ using a fluorescence scanner or recovered by reverse centrifugation for further analyses. Therefore, our chip possesses two functions: 1) it can be used for multi-target detection based on end-point in situ fluorescence detection; and 2) it can work as a sample preparation unit for analyses that need multiplex PCR such as hybridization and target sequencing. The performance of this chip was carefully examined and further illustrated in the identification of 8 pathogenic bacterial genomic DNA samples and 13 drug-resistance genes. Due to simplicity of its structure and operation, accuracy and generality, high-throughput capacity, and versatile functions (i.e., for in situ detection and sample preparation), our multiplex PCR chip has great potential in clinical diagnostics and nucleic acid-based point-of-care testing.

  8. Packaging commercial CMOS chips for lab on a chip integration.

    PubMed

    Datta-Chaudhuri, Timir; Abshire, Pamela; Smela, Elisabeth

    2014-05-21

    Combining integrated circuitry with microfluidics enables lab-on-a-chip (LOC) devices to perform sensing, freeing them from benchtop equipment. However, this integration is challenging with small chips, as is briefly reviewed with reference to key metrics for package comparison. In this paper we present a simple packaging method for including mm-sized, foundry-fabricated dies containing complementary metal oxide semiconductor (CMOS) circuits within LOCs. The chip is embedded in an epoxy handle wafer to yield a level, large-area surface, allowing subsequent photolithographic post-processing and microfluidic integration. Electrical connection off-chip is provided by thin film metal traces passivated with parylene-C. The parylene is patterned to selectively expose the active sensing area of the chip, allowing direct interaction with a fluidic environment. The method accommodates any die size and automatically levels the die and handle wafer surfaces. Functionality was demonstrated by packaging two different types of CMOS sensor ICs, a bioamplifier chip with an array of surface electrodes connected to internal amplifiers for recording extracellular electrical signals and a capacitance sensor chip for monitoring cell adhesion and viability. Cells were cultured on the surface of both types of chips, and data were acquired using a PC. Long term culture (weeks) showed the packaging materials to be biocompatible. Package lifetime was demonstrated by exposure to fluids over a longer duration (months), and the package was robust enough to allow repeated sterilization and re-use. The ease of fabrication and good performance of this packaging method should allow wide adoption, thereby spurring advances in miniaturized sensing systems. PMID:24682025

  9. A highly efficient and effective motif discovery method for ChIP-seq/ChIP-chip data using positional information

    PubMed Central

    Ma, Xiaotu; Kulkarni, Ashwinikumar; Zhang, Zhihua; Xuan, Zhenyu; Serfling, Robert; Zhang, Michael Q.

    2012-01-01

    Identification of DNA motifs from ChIP-seq/ChIP-chip [chromatin immunoprecipitation (ChIP)] data is a powerful method for understanding the transcriptional regulatory network. However, most established methods are designed for small sample sizes and are inefficient for ChIP data. Here we propose a new k-mer occurrence model to reflect the fact that functional DNA k-mers often cluster around ChIP peak summits. With this model, we introduced a new measure to discover functional k-mers. Using simulation, we demonstrated that our method is more robust against noises in ChIP data than available methods. A novel word clustering method is also implemented to group similar k-mers into position weight matrices (PWMs). Our method was applied to a diverse set of ChIP experiments to demonstrate its high sensitivity and specificity. Importantly, our method is much faster than several other methods for large sample sizes. Thus, we have developed an efficient and effective motif discovery method for ChIP experiments. PMID:22228832

  10. ChIP bias as a function of cross-linking time.

    PubMed

    Baranello, Laura; Kouzine, Fedor; Sanford, Suzanne; Levens, David

    2016-05-01

    The chromatin immunoprecipitation (ChIP) assay is widely used to capture interactions between chromatin and regulatory proteins in vivo. Formaldehyde cross-linking of DNA and proteins is a critical step required to trap their interactions inside the cells before immunoprecipitation and analysis. Yet insufficient attention has been given to variables that might give rise to artifacts in this procedure, such as the duration of cross-linking. We analyzed the dependence of the ChIP signal on the duration of formaldehyde cross-linking time for two proteins: DNA topoisomerase 1 (Top1) that is functionally associated with the double helix in vivo, especially with active chromatin, and green fluorescent protein (GFP) that has no known bona fide interactions with DNA. With short time of formaldehyde fixation, only Top1 immunoprecipation efficiently recovered DNA from active promoters, whereas prolonged fixation augmented non-specific recovery of GFP dramatizing the need to optimize ChIP protocols to minimize the time of cross-linking, especially for abundant nuclear proteins. Thus, ChIP is a powerful approach to study the localization of protein on the genome when care is taken to manage potential artifacts. PMID:26685864

  11. Microfluidic Chips for Semen Analysis

    PubMed Central

    Segerink, L.I.; Sprenkels, A.J.; Oosterhuis, G.J.E.; Vermes, I.; van den Berg, A.

    2012-01-01

    The gold standard of semen analysis is still an manual method, which is time-consuming, labour intensive and needs thorough quality control. Microfluidics can also offer advantages for this application. Therefore a first step in the development of a microfluidic chip has been made, which enables the man the semen analysis at home. In this article recent efforts to determine the concentration and motility using a microfluidic chip are summarized.

  12. Rutger's CAM2000 chip architecture

    NASA Technical Reports Server (NTRS)

    Smith, Donald E.; Hall, J. Storrs; Miyake, Keith

    1993-01-01

    This report describes the architecture and instruction set of the Rutgers CAM2000 memory chip. The CAM2000 combines features of Associative Processing (AP), Content Addressable Memory (CAM), and Dynamic Random Access Memory (DRAM) in a single chip package that is not only DRAM compatible but capable of applying simple massively parallel operations to memory. This document reflects the current status of the CAM2000 architecture and is continually updated to reflect the current state of the architecture and instruction set.

  13. Application of the chromatin immunoprecipitation method to identify in vivo protein-DNA associations in fission yeast.

    PubMed

    Takahashi, K; Saitoh, S; Yanagida, M

    2000-10-31

    The chromatin immunoprecipitation (ChIP) method provides an ideal tool for detecting direct or indirect interactions between proteins of interest and DNAs with known sequences. Here, we introduce the ChIP protocol used in our laboratory to identify in vivo protein-DNA association in the fission yeast Schizosaccharomyces pombe. The cytological and genetic merits of the fission yeast for studying control of the eukaryotic cell cycle and chromosome dynamics are reinforced by application of this ChIP method.

  14. CRRES microelectronic test chip

    NASA Astrophysics Data System (ADS)

    Lin, Y.-S.; Buehler, M. G.; Ray, K. P.; Sokoloski, M. M.

    1991-12-01

    The JPL CRRES chip was designed and fabricated in 1985 and included in the CRRES MEP. MOSFET Matrix results show the effect of shielding on radiation-induced MOSFET threshold voltage shifts and channel mobility degradation. Shielded (middle board) MOSFETs have a threshold-voltage damage factor that is approximately three orders of magnitude smaller than would be estimated from Co-60 ground tests. Unshielded (outer board) MOSFETs have a threshold-voltage damage factor that would be estimated from Co-60 ground tests. Temperature swings as large as 23 C with a 22.5 orbit periodicity affected the MOSFET data and were removed from the data in order to reveal the radiation effects. This experiment demonstrated the feasibility of characterizing MOSFETs in a matrix, thus reducing the complexity and mass of the experiment.

  15. Analysis Methods of Magnesium Chips

    NASA Astrophysics Data System (ADS)

    Ohmann, Sven; Ditze, André; Scharf, Christiane

    2015-11-01

    The quality of recycled magnesium from chips depends strongly on their exposure to inorganic and organic impurities that are added during the production processes. Different kinds of magnesium chips from these processes were analyzed by several methods. In addition, the accuracy and effectiveness of the methods are discussed. The results show that the chips belong either to the AZ91, AZ31, AM50/60, or AJ62 alloy. Some kinds of chips show deviations from the above-mentioned normations. Different impurities result mainly from transition metals and lime. The water and oil content does not exceed 25%, and the chip size is not more than 4 mm in the diameter. The sieve analysis shows good results for oily and wet chips. The determination of oil and water shows better results for the application of a Soxhlet compared with the addition of lime and vacuum distillation. The most accurate values for the determination of water and oil are obtained by drying at 110°C (for water) and washing with acetone (for oil) by hand.

  16. GeoChips for Analysis of Microbial Functional Communities

    SciTech Connect

    Van Nostrand, Joy D.; Wu, Liyou; He, Zhili; Zhou, Jizhong

    2008-09-30

    Functional gene arrays (FGA) are microarrays that contain probes for genes encoding proteins or enzymes involved in functions of interest and allow for the study of thousands of genes at one time. The most comprehensive FGA to date is the GeoChip, which contains ~;;24,000 probes for ~;;10,000 genes involved in the geochemical cycling of C, N, P, and S, as well as genes involved in metal resistance and reduction and contaminant degradation. This chapter details the methods necessary for GeoChip analysis. Methods covered include preparation of DNA (whole community genome amplification and labeling), array setup (prehybridization steps), hybridization (sample and hybridization buffers), and post hybridization steps (slide washing and array scanning).

  17. Stem end chip defect in tubers used for potato chip production

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Stem-end chip defect (SECD) is a serious tuber quality concern that affects chipping potatoes (Solanum tuberosum). SECD defect is characterized by dark-colored vascular tissues and adjacent cortical tissues at the tuber stem-end portion of potato chips after frying. Chips with SECD are unattractive ...

  18. Process for 3D chip stacking

    DOEpatents

    Malba, V.

    1998-11-10

    A manufacturable process for fabricating electrical interconnects which extend from a top surface of an integrated circuit chip to a sidewall of the chip using laser pantography to pattern three dimensional interconnects. The electrical interconnects may be of an L-connect or L-shaped type. The process implements three dimensional (3D) stacking by moving the conventional bond or interface pads on a chip to the sidewall of the chip. Implementation of the process includes: (1) holding individual chips for batch processing, (2) depositing a dielectric passivation layer on the top and sidewalls of the chips, (3) opening vias in the dielectric, (4) forming the interconnects by laser pantography, and (5) removing the chips from the holding means. The process enables low cost manufacturing of chips with bond pads on the sidewalls, which enables stacking for increased performance, reduced space, and higher functional per unit volume. 3 figs.

  19. Process for 3D chip stacking

    DOEpatents

    Malba, Vincent

    1998-01-01

    A manufacturable process for fabricating electrical interconnects which extend from a top surface of an integrated circuit chip to a sidewall of the chip using laser pantography to pattern three dimensional interconnects. The electrical interconnects may be of an L-connect or L-shaped type. The process implements three dimensional (3D) stacking by moving the conventional bond or interface pads on a chip to the sidewall of the chip. Implementation of the process includes: 1) holding individual chips for batch processing, 2) depositing a dielectric passivation layer on the top and sidewalls of the chips, 3) opening vias in the dielectric, 4) forming the interconnects by laser pantography, and 5) removing the chips from the holding means. The process enables low cost manufacturing of chips with bond pads on the sidewalls, which enables stacking for increased performance, reduced space, and higher functional per unit volume.

  20. Microfluidic interface technology based on stereolithography for glass-based lab-on-a-chips.

    PubMed

    Han, Song-I; Han, Ki-Ho

    2013-01-01

    As lab-on-a-chips are developed for on-chip integrated microfluidic systems with multiple functions, the development of microfluidic interface (MFI) technology to enable integration of complex microfluidic systems becomes increasingly important and faces many technical difficulties. Such difficulties include the need for more complex structures, the possibility of biological or chemical cross-contamination between functional compartments, and the possible need for individual compartments fabricated from different substrate materials. This chapter introduces MFI technology, based on rapid stereolithography, for a glass-based miniaturized genetic sample preparation system, as an example of a complex lab-on-a-chip that could include functional elements such as; solid-phase DNA extraction, polymerase chain reaction, and capillary electrophoresis. To enable the integration of a complex lab-on-a-chip system in a single chip, MFI technology based on stereolithography provides a simple method for realizing complex arrangements of one-step plug-in microfluidic interconnects, integrated microvalves for microfluidic control, and optical windows for on-chip optical processes.

  1. Lab-on-a-Chip

    NASA Technical Reports Server (NTRS)

    2004-01-01

    Labs on chips are manufactured in many shapes and sizes and can be used for numerous applications, from medical tests to water quality monitoring to detecting the signatures of life on other planets. The eight holes on this chip are actually ports that can be filled with fluids or chemicals. Tiny valves control the chemical processes by mixing fluids that move in the tiny channels that look like lines, connecting the ports. Scientists at NASA's Marshall Space Flight Center (MSFC) in Huntsville, Alabama designed this chip to grow biological crystals on the International Space Station. Through this research, they discovered that this technology is ideally suited for solving the challenges of the Vision for Space Exploration. For example, thousands of chips the size of dimes could be loaded on a Martian rover looking for biosignatures of past or present life. Other types of chips could be placed in handheld devices used to monitor microbes in water or to quickly conduct medical tests on astronauts. (NASA/MSFC/D.Stoffer)

  2. Detection of K-Ras oncogene using magnetic beads-quantum dots in microfluidic chip.

    PubMed

    Noh, Han Na; Kim, Jong Sung

    2013-08-01

    Recently quantum dots (QDs) have been extensively used in the field of biotechnology. QDs have merits of wide selection of emission wavelength and exceptional stability against photo bleaching over conventional organic fluorophores and are used in cell imaging, biomarker, and fluorescence resonance energy transfer (FRET) sensor. Magnetic beads have been used as solid support in microfluidic devices to trace bio-molecules. In this study, Polydimethylsiloxane (PDMS) based microfluidic chips were prepared for the detection of K-Ras oncogene by using QDs-DNA conjugate. K-Ras oncogene can be detected by fluorescence quenching in microfluidic chip. Carboxylated CdSe/ZnS QDs (emission wavelength: 605 nm) could bind to magnetic beads of polystyrene/divinyl benzene via EDC/NHS crosslinking reaction. The fluorescence from QDs could be quenched by intercalating dye (thiazol orange dimers: TOTO-3) after hybridization with target DNA and probe DNA in the channel of microfluidic chip. The fluorescence intensity change of QDs after hybridization in microfluidic chip has been studied. PMID:23882748

  3. Growth of immobilized DNA by polymerase: bridging nanoelectrodes with individual dsDNA molecules.

    PubMed

    Linko, Veikko; Leppiniemi, Jenni; Shen, Boxuan; Niskanen, Einari; Hytönen, Vesa P; Toppari, J Jussi

    2011-09-01

    We present a method for controlled connection of gold electrodes with dsDNA molecules (locally on a chip) by utilizing polymerase to elongate single-stranded DNA primers attached to the electrodes. Thiol-modified oligonucleotides are directed and immobilized to nanoscale electrodes by means of dielectrophoretic trapping, and extended in a procedure mimicking PCR, finally forming a complete dsDNA molecule bridging the gap between the electrodes. The technique opens up opportunities for building from the bottom-up, for detection and sensing applications, and also for molecular electronics.

  4. Growth of immobilized DNA by polymerase: bridging nanoelectrodes with individual dsDNA molecules

    NASA Astrophysics Data System (ADS)

    Linko, Veikko; Leppiniemi, Jenni; Shen, Boxuan; Niskanen, Einari; Hytönen, Vesa P.; Toppari, J. Jussi

    2011-09-01

    We present a method for controlled connection of gold electrodes with dsDNA molecules (locally on a chip) by utilizing polymerase to elongate single-stranded DNA primers attached to the electrodes. Thiol-modified oligonucleotides are directed and immobilized to nanoscale electrodes by means of dielectrophoretic trapping, and extended in a procedure mimicking PCR, finally forming a complete dsDNA molecule bridging the gap between the electrodes. The technique opens up opportunities for building from the bottom-up, for detection and sensing applications, and also for molecular electronics.

  5. Camera-on-a-Chip

    NASA Technical Reports Server (NTRS)

    1999-01-01

    Jet Propulsion Laboratory's research on a second generation, solid-state image sensor technology has resulted in the Complementary Metal- Oxide Semiconductor Active Pixel Sensor (CMOS), establishing an alternative to the Charged Coupled Device (CCD). Photobit Corporation, the leading supplier of CMOS image sensors, has commercialized two products of their own based on this technology: the PB-100 and PB-300. These devices are cameras on a chip, combining all camera functions. CMOS "active-pixel" digital image sensors offer several advantages over CCDs, a technology used in video and still-camera applications for 30 years. The CMOS sensors draw less energy, they use the same manufacturing platform as most microprocessors and memory chips, and they allow on-chip programming of frame size, exposure, and other parameters.

  6. Beyond the Gene Chip

    PubMed Central

    Heng, J. B; Aksimentiev, A.; Ho, C.; Dimitrov, V.; Sorsch, T.; Miner, J.; Mansfield, W.; Schulten, K.; Timp, G.

    2008-01-01

    We describe a prospective strategy for reading the encyclopedic information encoded in the genome: using a nanopore in a membrane formed from an MOS-capacitor to sense the charge in DNA. In principle, as DNA permeates the capacitor-membrane through the pore, the electrostatic charge distribution characteristic of the molecule should polarize the capacitor and induce a voltage on the electrodes that can be measured. Silicon nanofabrication and molecular dynamic simulations with atomic detail are technological linchpins in the development of this detector. The sub-nanometer precision available through silicon nanotechnology facilitates the fabrication of the detector, and molecular dynamics provides us with a means to design it and analyze the experimental outcomes. PMID:18815623

  7. Localization and functional analysis of CHIP28k water channels in stably transfected Chinese hamster ovary cells.

    PubMed

    Ma, T; Frigeri, A; Tsai, S T; Verbavatz, J M; Verkman, A S

    1993-10-25

    CHIP28 is a major water transporting protein in erythrocytes and plasma membranes in kidney proximal tubule and thin descending limb of Henle. Chinese hamster ovary cells were stably transfected with the coding sequence of cloned rat kidney CHIP28k using expression vectors containing cytomegalovirus or Rous sarcoma virus promoters. Clonal cell populations expressed a 1.3-kilobase mRNA on Northern blot probed by CHIP28k cDNA and a 28-kDa protein on immunoblot probed by a polyclonal CHIP28 antibody. The clone with greatest expression produced approximately 8 x 10(6) copies of CHIP28k protein/cell. Plasma membrane osmotic water permeability (Pf), measured by stopped-flow light scattering, was 0.004 cm/s in control (vector-transfected) cells (10 degrees C) and 0.014 cm/s in the CHIP28k-transfected cells. Pf in CHIP28k-transfected cells had an activation energy of 4.9 kcal/mol and was reversibly inhibited by HgCl2. CHIP28k expression did not affect the transport of protons and the small polar non-electrolytes urea and formamide. CHIP28k immunoreactivity and function was then determined in subcellular fractions. Pf in 6-carboxyfluorescein-labeled endocytic vesicles, measured by a stopped-flow fluorescence quenching assay, was 0.002 cm/s (control cells) and 0.011 cm/s (CHIP28k-transfected cells); Pf in transfected cells was inhibited by HgCl2. Immunoblotting of fractionated endoplasmic reticulum, Golgi, and plasma membranes revealed high densities of CHIP28k (approximately 5000 monomers/microns 2 in plasma membrane) with different glycosylation patterns; functional water transport activity was present only in Golgi and plasma membrane vesicles. Antibody detection of CHIP28k by confocal fluorescence microscopy and immunogold electron microscopy revealed localization to plasma membrane and intracellular vesicles. These studies establish a stably transfected somatic cell line that strongly expresses functional CHIP28k water channels. As in the original proximal tubule cells

  8. Single molecule actuation and detection on a lab-on-a-chip magnetoresistive platform

    NASA Astrophysics Data System (ADS)

    Chaves, R. C.; Bensimon, D.; Freitas, P. P.

    2011-03-01

    On-chip magnetic tweezers based on current loops were integrated with magnetoresistive sensors. Magnetic forces up to 1.0±0.3pN are produced to actuate on DNA anchored to the surface of a flow cell and labeled with micrometer-sized magnetic beads. The levitation of the beads stretches the immobilized DNA. The relative position of the magnetic beads is monitored using spin-valve sensors. A bead vertical displacement resolution of 60nm is derived for DNA molecular motor activity in a tweezer steady current regime.

  9. Chip Advancer For GPS Receiver

    NASA Technical Reports Server (NTRS)

    Meehan, Thomas K.; Srinivasan, Jeffrey M.; Thomas, J. Brooks

    1989-01-01

    Instrument errors made negligible. For each integration interval, both delay and rate of change of delay initialized to small fraction of chip - for example, to order of 10 to the negative 7th power - thereby making feedback control and extraction of delay highly accurate and flexible. With appropriate selection of sampling rate relative to chip rate, commensurability errors reduced to extremely small levels. In Global Positioning System (GPS) receiver, pseudorandom code sequence generated by simple digital logic incorporating effects of time, delay, and rate of change of delay. Flexibility in starting time and sum interval very useful in aligning correlation interval with beginnings and endings of data bits.

  10. CMOS foveal image sensor chip

    NASA Technical Reports Server (NTRS)

    Bandera, Cesar (Inventor); Scott, Peter (Inventor); Sridhar, Ramalingam (Inventor); Xia, Shu (Inventor)

    2002-01-01

    A foveal image sensor integrated circuit comprising a plurality of CMOS active pixel sensors arranged both within and about a central fovea region of the chip. The pixels in the central fovea region have a smaller size than the pixels arranged in peripheral rings about the central region. A new photocharge normalization scheme and associated circuitry normalizes the output signals from the different size pixels in the array. The pixels are assembled into a multi-resolution rectilinear foveal image sensor chip using a novel access scheme to reduce the number of analog RAM cells needed. Localized spatial resolution declines monotonically with offset from the imager's optical axis, analogous to biological foveal vision.

  11. Programmable Multi-Chip Module

    DOEpatents

    Kautz, David; Morgenstern, Howard; Blazek, Roy J.

    2004-11-16

    A multi-chip module comprising a low-temperature co-fired ceramic substrate having a first side on which are mounted active components and a second side on which are mounted passive components, wherein this segregation of components allows for hermetically sealing the active components with a cover while leaving accessible the passive components, and wherein the passive components are secured using a reflow soldering technique and are removable and replaceable so as to make the multi-chip module substantially programmable with regard to the passive components.

  12. Programmable multi-chip module

    DOEpatents

    Kautz, David; Morgenstern, Howard; Blazek, Roy J.

    2004-03-02

    A multi-chip module comprising a low-temperature co-fired ceramic substrate having a first side on which are mounted active components and a second side on which are mounted passive components, wherein this segregation of components allows for hermetically sealing the active components with a cover while leaving accessible the passive components, and wherein the passive components are secured using a reflow soldering technique and are removable and replaceable so as to make the multi-chip module substantially programmable with regard to the passive components.

  13. Programmable Multi-Chip Module

    DOEpatents

    Kautz, David; Morgenstern, Howard; Blazek, Roy J.

    2005-05-24

    A multi-chip module comprising a low-temperature co-fired ceramic substrate having a first side on which are mounted active components and a second side on which are mounted passive components, wherein this segregation of components allows for hermetically sealing the active components with a cover while leaving accessible the passive components, and wherein the passive components are secured using a reflow soldering technique and are removable and replaceable so as to make the multi-chip module substantially programmable with regard to the passive components.

  14. Mapping Protein-DNA Interactions Using ChIP-exo and Illumina-Based Sequencing.

    PubMed

    Barfeld, Stefan J; Mills, Ian G

    2016-01-01

    Chromatin immunoprecipitation (ChIP) provides a means of enriching DNA associated with transcription factors, histone modifications, and indeed any other proteins for which suitably characterized antibodies are available. Over the years, sequence detection has progressed from quantitative real-time PCR and Southern blotting to microarrays (ChIP-chip) and now high-throughput sequencing (ChIP-seq). This progression has vastly increased the sequence coverage and data volumes generated. This in turn has enabled informaticians to predict the identity of multi-protein complexes on DNA based on the overrepresentation of sequence motifs in DNA enriched by ChIP with a single antibody against a single protein. In the course of the development of high-throughput sequencing, little has changed in the ChIP methodology until recently. In the last three years, a number of modifications have been made to the ChIP protocol with the goal of enhancing the sensitivity of the method and further reducing the levels of nonspecific background sequences in ChIPped samples. In this chapter, we provide a brief commentary on these methodological changes and describe a detailed ChIP-exo method able to generate narrower peaks and greater peak coverage from ChIPped material.

  15. Atom chip gravimeter

    NASA Astrophysics Data System (ADS)

    Schubert, Christian; Abend, Sven; Gebbe, Martina; Gersemann, Matthias; Ahlers, Holger; Müntinga, Hauke; Matthias, Jonas; Sahelgozin, Maral; Herr, Waldemar; Lämmerzahl, Claus; Ertmer, Wolfgang; Rasel, Ernst

    2016-04-01

    Atom interferometry has developed into a tool for measuring rotations [1], accelerations [2], and testing fundamental physics [3]. Gravimeters based on laser cooled atoms demonstrated residual uncertainties of few microgal [2,4] and were simplified for field applications [5]. Atomic gravimeters rely on the interference of matter waves which are coherently manipulated by laser light fields. The latter can be interpreted as rulers to which the position of the atoms is compared. At three points in time separated by a free evolution, the light fields are pulsed onto the atoms. First, a coherent superposition of two momentum states is produced, then the momentum is inverted, and finally the two trajectories are recombined. Depending on the acceleration the atoms experienced, the number of atoms detected in the output ports will change. Consequently, the acceleration can be determined from the output signal. The laser cooled atoms with microkelvin temperatures used in state-of-the-art gravimeters impose limits on the accuracy [4]. Therefore, ultra-cold atoms generated by Bose-Einstein condensation and delta-kick collimation [6,7] are expected to be the key for further improvements. These sources suffered from a low flux implying an incompatible noise floor, but a competitive performance was demonstrated recently with atom chips [8]. In the compact and robust setup constructed for operation in the drop tower [6] we demonstrated all steps necessary for an atom chip gravimeter with Bose-Einstein condensates in a ground based operation. We will discuss the principle of operation, the current performance, and the perspectives to supersede the state of the art. The authors thank the QUANTUS cooperation for contributions to the drop tower project in the earlier stages. This work is supported by the German Space Agency (DLR) with funds provided by the Federal Ministry for Economic Affairs and Energy (BMWi) due to an enactment of the German Bundestag under grant numbers DLR 50WM

  16. Protein Chips for Detection of Salmonella spp. from Enrichment Culture.

    PubMed

    Poltronieri, Palmiro; Cimaglia, Fabio; De Lorenzis, Enrico; Chiesa, Maurizio; Mezzolla, Valeria; Reca, Ida Barbara

    2016-01-01

    Food pathogens are the cause of foodborne epidemics, therefore there is a need to detect the pathogens in food productions rapidly. A pre-enrichment culture followed by selective agar plating are standard detection methods. Molecular methods such as qPCR have provided a first rapid protocol for detection of pathogens within 24 h of enrichment culture. Biosensors also may provide a rapid tool to individuate a source of Salmonella contamination at early times of pre-enrichment culture. Forty mL of Salmonella spp. enrichment culture were processed by immunoseparation using the Pathatrix, as in AFNOR validated qPCR protocols. The Salmonella biosensor combined with immunoseparation showed a limit of detection of 100 bacteria/40 mL, with a 400 fold increase to previous results. qPCR analysis requires processing of bead-bound bacteria with lysis buffer and DNA clean up, with a limit of detection of 2 cfu/50 μL. Finally, a protein chip was developed and tested in screening and identification of 5 common pathogen species, Salmonella spp., E. coli, S. aureus, Campylobacter spp. and Listeria spp. The protein chip, with high specificity in species identification, is proposed to be integrated into a Lab-on-Chip system, for rapid and reproducible screening of Salmonella spp. and other pathogen species contaminating food productions. PMID:27110786

  17. Protein Chips for Detection of Salmonella spp. from Enrichment Culture

    PubMed Central

    Poltronieri, Palmiro; Cimaglia, Fabio; De Lorenzis, Enrico; Chiesa, Maurizio; Mezzolla, Valeria; Reca, Ida Barbara

    2016-01-01

    Food pathogens are the cause of foodborne epidemics, therefore there is a need to detect the pathogens in food productions rapidly. A pre-enrichment culture followed by selective agar plating are standard detection methods. Molecular methods such as qPCR have provided a first rapid protocol for detection of pathogens within 24 h of enrichment culture. Biosensors also may provide a rapid tool to individuate a source of Salmonella contamination at early times of pre-enrichment culture. Forty mL of Salmonella spp. enrichment culture were processed by immunoseparation using the Pathatrix, as in AFNOR validated qPCR protocols. The Salmonella biosensor combined with immunoseparation showed a limit of detection of 100 bacteria/40 mL, with a 400 fold increase to previous results. qPCR analysis requires processing of bead-bound bacteria with lysis buffer and DNA clean up, with a limit of detection of 2 cfu/50 μL. Finally, a protein chip was developed and tested in screening and identification of 5 common pathogen species, Salmonella spp., E. coli, S. aureus, Campylobacter spp. and Listeria spp. The protein chip, with high specificity in species identification, is proposed to be integrated into a Lab-on-Chip system, for rapid and reproducible screening of Salmonella spp. and other pathogen species contaminating food productions. PMID:27110786

  18. Recovery Based Nanowire Field-Effect Transistor Detection of Pathogenic Avian Influenza DNA

    NASA Astrophysics Data System (ADS)

    Lin, Chih-Heng; Chu, Chia-Jung; Teng, Kang-Ning; Su, Yi-Jr; Chen, Chii-Dong; Tsai, Li-Chu; Yang, Yuh-Shyong

    2012-02-01

    Fast and accurate diagnosis is critical in infectious disease surveillance and management. We proposed a DNA recovery system that can easily be adapted to DNA chip or DNA biosensor for fast identification and confirmation of target DNA. This method was based on the re-hybridization of DNA target with a recovery DNA to free the DNA probe. Functionalized silicon nanowire field-effect transistor (SiNW FET) was demonstrated to monitor such specific DNA-DNA interaction using high pathogenic strain virus hemagglutinin 1 (H1) DNA of avian influenza (AI) as target. Specific electric changes were observed in real-time for AI virus DNA sensing and device recovery when nanowire surface of SiNW FET was modified with complementary captured DNA probe. The recovery based SiNW FET biosensor can be further developed for fast identification and further confirmation of a variety of influenza virus strains and other infectious diseases.

  19. Trapping and manipulating single molecules of DNA

    NASA Astrophysics Data System (ADS)

    Shon, Min Ju

    This thesis presents the development and application of nanoscale techniques to trap and manipulate biomolecules, with a focus on DNA. These methods combine single-molecule microscopy and nano- and micro-fabrication to study biophysical properties of DNA and proteins. The Dimple Machine is a lab-on-a-chip device that can isolate and confine a small number of molecules from a bulk solution. It traps molecules in nanofabricated chambers, or "dimples", and the trapped molecules are then studied on a fluorescence microscope at the single-molecule level. The sampling of bulk solution by dimples is representative, reproducible, and automated, enabling highthroughput single-molecule experiments. The device was applied to study hybridization of oligonucleotides, particularly in the context of reaction thermodynamics and kinetics in nanoconfinement. The DNA Pulley is a system to study protein binding and the local mechanical properties of DNA. A molecule of DNA is tethered to a surface on one end, and a superparamagnetic bead is attached to the other. A magnet pulls the DNA taut, and a silicon nitride knife with a nanoscale blade scans the DNA along its contour. Information on the local properties of the DNA is extracted by tracking the bead with nanometer precision in a white-light microscope. The system can detect proteins bound to DNA and localize their recognition sites, as shown with a model protein, EcoRI restriction enzyme. Progress on the measurements of nano-mechanical properties of DNA is included.

  20. Thermodynamics and Kinetics of DNA Nanostructure Assembly

    NASA Astrophysics Data System (ADS)

    Nangreave, Jeanette

    2011-12-01

    The unique structural features of deoxyribonucleic acid (DNA) that are of considerable biological interest also make it a valuable engineering material. Perhaps the most useful property of DNA for molecular engineering is its ability to self-assemble into predictable, double helical secondary structures. These interactions are exploited to design a variety of DNA nanostructures, which can be organized into both discrete and periodic structures. This dissertation focuses on studying the dynamic behavior of DNA nanostructure recognition processes. The thermodynamics and kinetics of nanostructure binding are evaluated, with the intention of improving our ability to understand and control their assembly. Presented here are a series of studies toward this goal. First, multi-helical DNA nanostructures were used to investigate how the valency and arrangement of the connections between DNA nanostructures affect super-structure formation. The study revealed that both the number and the relative position of connections play a significant role in the stability of the final assembly. Next, several DNA nanostructures were designed to gain insight into how small changes to the nanostructure scaffolds, intended to vary their conformational flexibility, would affect their association equilibrium. This approach yielded quantitative information about the roles of enthalpy and entropy in the affinity of polyvalent DNA nanostructure interactions, which exhibit an intriguing compensating effect. Finally, a multi-helical DNA nanostructure was used as a model 'chip' for the detection of a single stranded DNA target. The results revealed that the rate constant of hybridization is strongly dominated by a rate-limiting nucleation step.

  1. Mitochondrial DNA.

    ERIC Educational Resources Information Center

    Wright, Russell G.; Bottino, Paul J.

    1986-01-01

    Provides background information for teachers on mitochondrial DNA, pointing out that it may have once been a free-living organism. Includes a ready-to-duplicate exercise titled "Using Microchondrial DNA to Measure Evolutionary Distance." (JN)

  2. Chipped Paint Crater

    NASA Technical Reports Server (NTRS)

    2003-01-01

    [figure removed for brevity, see original site]

    Released 9 April 2003

    In the high northern latitudes NW of Alba Patera, a smooth mantle of material that covers the landscape appears chipped away from the rim of a large crater. The prominent scarp that has formed from the retreat of the mantle lacks the rounded appearance of other ice-rich mantles found in the mid-latitudes. The nature of this mantling layer therefore is more enigmatic.

    Note: this THEMIS visual image has not been radiometrically nor geometrically calibrated for this preliminary release. An empirical correction has been performed to remove instrumental effects. A linear shift has been applied in the cross-track and down-track direction to approximate spacecraft and planetary motion. Fully calibrated and geometrically projected images will be released through the Planetary Data System in accordance with Project policies at a later time.

    NASA's Jet Propulsion Laboratory manages the 2001 Mars Odyssey mission for NASA's Office of Space Science, Washington, D.C. The Thermal Emission Imaging System (THEMIS) was developed by Arizona State University, Tempe, in collaboration with Raytheon Santa Barbara Remote Sensing. The THEMIS investigation is led by Dr. Philip Christensen at Arizona State University. Lockheed Martin Astronautics, Denver, is the prime contractor for the Odyssey project, and developed and built the orbiter. Mission operations are conducted jointly from Lockheed Martin and from JPL, a division of the California Institute of Technology in Pasadena.

    Image information: VIS instrument. Latitude 62.9, Longitude 226.2 East (133.8 West). 19 meter/pixel resolution.

  3. Wireless Charge Based Capacitance Measurement Circuits with On-Chip Spiral Inductor for Radio Frequency Identification Biosensor

    NASA Astrophysics Data System (ADS)

    Kim, Boram; Uno, Shigeyasu; Nakazato, Kazuo

    2012-04-01

    A wireless measuring system of charge based capacitance measurement (CBCM) circuit has been designed and demonstrated for biomedical applications. The radio frequency identification (RFID) chip that includes on-chip spiral inductor tag antenna, and RFID circuit, and CBCM sensor chip are fabricated within standard complementary metal oxide semiconductor (CMOS) process. The capacitance change caused by DNA detection can be converted into the voltage output using capacitance-to-voltage conversion circuit. To confirm the transmission of the capacitance, the poly-capacitor of fixed capacitance and on-chip spiral inductor tag antenna were fabricated using 1.2 µm, 2-metal, 2-poly CMOS technology. As a result of measurement, three different capacitances (34, 141, 564 fF) were detected wirelessly.

  4. DNA Banking

    SciTech Connect

    Reilly, P.R. )

    1992-11-01

    The author is involved in the ethical, legal, and social issues of banking of DNA and data from DNA analysis. In his attempt to determine the extent of DNA banking in the U.S., the author surveyed some commercial companies performing DNA banking services. This article summarizes the results of that survey, with special emphasis on the procedures the companies use to protect the privacy of individuals. 4 refs.

  5. Beyond the dna: a prototype for functional genomics

    SciTech Connect

    Albala, J

    2000-03-02

    A prototype oligonucleotide ''functional chip'' has been developed to screen novel DNA repair proteins for their ability to bind or alter different forms of DNA. This chip has been developed as a functional genomics screen for analysis of protein-DNA interactions for novel proteins identified from the Human Genome Project The process of novel gene identification that has ensued as a consequence of available sequence information is remarkable. The challenge how lies in determining the function of newly identified gene products in a time-and cost-effective high-throughput manner. The functional chip is generated by the robotic application of DNA spotted in a microarray format onto a glass slide. Individual proteins are then analyzed against the different form of DNA bound to the slide. Several prototype functional chips were designed to contain various DNA fragments tethered to a glass slide for analysis of protein-DNA binding or enzymatic activity of known proteins. The technology has been developed to screen novel, putative DNA repair proteins for their ability to bind various types of DNA alone and in concert with protein partners. An additional scheme has been devised to screen putative repair enzymes for their ability to process different types of DNA molecules. Current methods to analyze gene expression primarily utilize either of two technologies. The oligonucleotide chip, pioneered by Fodor and co-workers and Affymetrix, Inc., consists of greater than 64,000 oligonucleotides attached in situ to a glass support. The oligonucleotide chip has been used primarily to identify specific mutations in a given gene by hybridization against a fluorescently-labeled substrate. The second method is the microarray, whereby DNA targets are systematically arranged on a glass slide and then hybridized with fluorescently-labeled complex targets for gene expression analysis (Jordan, 1998). By this technique, a large amount of information can be obtained examining global

  6. An integrated microfluidic chip for the detection of bacteria - A proof of concept.

    PubMed

    Guo, Zhe; Yu, Ting; He, Jiarui; Liu, Fen; Hao, Hualong; Zhao, Yang; Wen, Jiabin; Wang, Qi

    2015-08-01

    We designed a microfluidic chip as a proof of concept for the detection of bacterial DNA. The chip was fabricated with poly-dimethylsiloxane (PDMS). It included a solid phase extraction (SPE) chamber, two separate channels and multiple loop-mediated isothermal amplification (LAMP) chambers. Three bacterial strains (Escherichia coli O157:H7, methicillin-resistant Staphylococcus aureus and methicillin-sensitive S. aureus) were used to test the feasibility of the device. LAMP products were examined directly using a UV light and verified by agarose gel electrophoresis. Using this chip, we successfully detected E. coli O157:H7, MSSA and MRSA in less than 2 h. The detection limit for genes rfbE, spa and mecA (specific to E. coli O157:H7, MSSA and MRSA, respectively) was <10(2) CFU/100 μl. Further work is required to refine this approach and rigorously assess its analytical and diagnostic specificity and sensitivity. PMID:25979593

  7. An integrated microfluidic chip for the detection of bacteria - A proof of concept.

    PubMed

    Guo, Zhe; Yu, Ting; He, Jiarui; Liu, Fen; Hao, Hualong; Zhao, Yang; Wen, Jiabin; Wang, Qi

    2015-08-01

    We designed a microfluidic chip as a proof of concept for the detection of bacterial DNA. The chip was fabricated with poly-dimethylsiloxane (PDMS). It included a solid phase extraction (SPE) chamber, two separate channels and multiple loop-mediated isothermal amplification (LAMP) chambers. Three bacterial strains (Escherichia coli O157:H7, methicillin-resistant Staphylococcus aureus and methicillin-sensitive S. aureus) were used to test the feasibility of the device. LAMP products were examined directly using a UV light and verified by agarose gel electrophoresis. Using this chip, we successfully detected E. coli O157:H7, MSSA and MRSA in less than 2 h. The detection limit for genes rfbE, spa and mecA (specific to E. coli O157:H7, MSSA and MRSA, respectively) was <10(2) CFU/100 μl. Further work is required to refine this approach and rigorously assess its analytical and diagnostic specificity and sensitivity.

  8. Detecting a single molecule using a micropore-nanopore hybrid chip

    PubMed Central

    2013-01-01

    Nanopore-based DNA sequencing and biomolecule sensing have attracted more and more attention. In this work, novel sensing devices were built on the basis of the chips containing nanopore arrays in polycarbonate (PC) membranes and micropores in Si3N4 films. Using the integrated chips, the transmembrane ionic current induced by biomolecule's translocation was recorded and analyzed, which suggested that the detected current did not change linearly as commonly expected with increasing biomolecule concentration. On the other hand, detailed translocation information (such as translocation gesture) was also extracted from the discrete current blockages in basic current curves. These results indicated that the nanofluidic device based on the chips integrated by micropores and nanopores possessed comparative potentials in biomolecule sensing. PMID:24261484

  9. Dna Sequencing

    DOEpatents

    Tabor, Stanley; Richardson, Charles C.

    1995-04-25

    A method for sequencing a strand of DNA, including the steps off: providing the strand of DNA; annealing the strand with a primer able to hybridize to the strand to give an annealed mixture; incubating the mixture with four deoxyribonucleoside triphosphates, a DNA polymerase, and at least three deoxyribonucleoside triphosphates in different amounts, under conditions in favoring primer extension to form nucleic acid fragments complementory to the DNA to be sequenced; labelling the nucleic and fragments; separating them and determining the position of the deoxyribonucleoside triphosphates by differences in the intensity of the labels, thereby to determine the DNA sequence.

  10. Molecular Sentinel-on-Chip for SERS-Based Biosensing†

    PubMed Central

    Du, Yan; Batchelor, Dale; Leonard, Donovan N.; Misra, Veena; Vo-Dinh, Tuan

    2013-01-01

    The development of DNA detection techniques on large-area plasmonics-active platforms is critical for many medical applications such as high-throughput screening, medical diagnosis and systems biology research. Here, we report for the first time a unique “molecular sentinel-on-chip” (MSC) technology for surface-enhanced Raman scattering (SERS)-based DNA detection. This unique approach allows label-free detection of DNA molecules on chips developed on a wafer scale using large area nanofabrication methodologies. To develop plasmonics-active biosensing platforms in a repeatable and reproducible manner, we employed a combination of deep UV lithography, atomic layer deposition, and metal deposition to fabricate triangular-shaped nanowire (TSNW) arrays having controlled sub-10 nm gaps nanostructures over an entire 6-inch wafer. The detection of a DNA sequence of the Ki-67 gene, a critical breast cancer biomarker, on the TSNW substrate illustrates the usefulness and potential of the MSC technology as a novel SERS-based DNA detection method. PMID:23493773

  11. Silicon ball grid array chip carrier

    DOEpatents

    Palmer, David W.; Gassman, Richard A.; Chu, Dahwey

    2000-01-01

    A ball-grid-array integrated circuit (IC) chip carrier formed from a silicon substrate is disclosed. The silicon ball-grid-array chip carrier is of particular use with ICs having peripheral bond pads which can be reconfigured to a ball-grid-array. The use of a semiconductor substrate such as silicon for forming the ball-grid-array chip carrier allows the chip carrier to be fabricated on an IC process line with, at least in part, standard IC processes. Additionally, the silicon chip carrier can include components such as transistors, resistors, capacitors, inductors and sensors to form a "smart" chip carrier which can provide added functionality and testability to one or more ICs mounted on the chip carrier. Types of functionality that can be provided on the "smart" chip carrier include boundary-scan cells, built-in test structures, signal conditioning circuitry, power conditioning circuitry, and a reconfiguration capability. The "smart" chip carrier can also be used to form specialized or application-specific ICs (ASICs) from conventional ICs. Types of sensors that can be included on the silicon ball-grid-array chip carrier include temperature sensors, pressure sensors, stress sensors, inertia or acceleration sensors, and/or chemical sensors. These sensors can be fabricated by IC processes and can include microelectromechanical (MEM) devices.

  12. Real-time PCR array chip with capillary-driven sample loading and reactor sealing for point-of-care applications.

    PubMed

    Ramalingam, Naveen; Liu, Hao-Bing; Dai, Chang-Chun; Jiang, Yu; Wang, Hui; Wang, Qinghui; M Hui, Kam; Gong, Hai-Qing

    2009-10-01

    A major challenge for the lab-on-a-chip (LOC) community is to develop point-of-care diagnostic chips that do not use instruments. Such instruments include pumping or liquid handling devices for distribution of patient's nucleic-acid test sample among an array of reactors and microvalves or mechanical parts to seal these reactors. In this paper, we report the development of a primer pair pre-loaded PCR array chip, in which the loading of the PCR mixture into an array of reactors and subsequent sealing of the reactors were realized by a novel capillary-based microfluidics with a manual two-step pipetting operations. The chip is capable of performing simultaneous (parallel) analyses of multiple gene targets and its performance was tested by amplifying twelve different gene targets against cDNA template from human hepatocellular carcinoma using SYBR Green I fluorescent dye. The versatility and reproducibility of the PCR-array chip are demonstrated by real-time PCR amplification of the BNI-1 fragment of SARS cDNA cloned in a plasmid vector. The reactor-to-reactor diffusion of the pre-loaded primer pairs in the chip is investigated to eliminate the possibility of primer cross-contamination. Key technical issues such as PCR mixture loss in gas-permeable PDMS chip layer and bubble generation due to different PDMS-glass bonding methods are investigated.

  13. Using DNA devices to track anticancer drug activity.

    PubMed

    Kahanda, Dimithree; Chakrabarti, Gaurab; Mcwilliams, Marc A; Boothman, David A; Slinker, Jason D

    2016-06-15

    It is beneficial to develop systems that reproduce complex reactions of biological systems while maintaining control over specific factors involved in such processes. We demonstrated a DNA device for following the repair of DNA damage produced by a redox-cycling anticancer drug, beta-lapachone (β-lap). These chips supported ß-lap-induced biological redox cycle and tracked subsequent DNA damage repair activity with redox-modified DNA monolayers on gold. We observed drug-specific changes in square wave voltammetry from these chips at therapeutic ß-lap concentrations of high statistical significance over drug-free control. We also demonstrated a high correlation of this change with the specific ß-lap-induced redox cycle using rational controls. The concentration dependence of ß-lap revealed significant signal changes at levels of high clinical significance as well as sensitivity to sub-lethal levels of ß-lap. Catalase, an enzyme decomposing peroxide, was found to suppress DNA damage at a NQO1/catalase ratio found in healthy cells, but was clearly overcome at a higher NQO1/catalase ratio consistent with cancer cells. We found that it was necessary to reproduce key features of the cellular environment to observe this activity. Thus, this chip-based platform enabled tracking of ß-lap-induced DNA damage repair when biological criteria were met, providing a unique synthetic platform for uncovering activity normally confined to inside cells. PMID:26901461

  14. DNA: Polymer and molecular code

    NASA Astrophysics Data System (ADS)

    Shivashankar, G. V.

    1999-10-01

    gene expression a prime example of a biological code. We developed a novel method of making DNA micro- arrays, the so-called DNA chip. Using the optical tweezer concept, we were able to pattern biomolecules on a solid substrate, developing a new type of sub-micron laser lithography. A laser beam is focused onto a thin gold film on a glass substrate. Laser ablation of gold results in local aggregation of nanometer scale beads conjugated with small DNA oligonucleotides, with sub-micron resolution. This leads to specific detection of cDNA and RNA molecules. We built a simple micro-array fabrication and detection in the laboratory, based on this method, to probe addressable pools (genes, proteins or antibodies). We have lately used molecular beacons (single stranded DNA with a stem-loop structure containing a fluorophore and quencher), for the direct detection of unlabelled mRNA. As a first step towards a study of the dynamics of the biological code, we have begun to examine the patterns of gene expression during virus (T7 phage) infection of E-coli bacteria.

  15. Genome-wide analysis of histone modifications by ChIP-chip to identify silenced genes in gastric cancer.

    PubMed

    Zhu, Xinjiang; Liu, Jian; Xu, Xiaoyang; Zhang, Chundong; Dai, Dongqiu

    2015-05-01

    The present study aimed to identify novel histone modification markers in gastric cancer (GC) by chromatin immunoprecipitation microarray (ChIP-chip) analysis and to determine whether these markers were able to discriminate between normal and GC cells. We also tested for correlations with DNA methylation. We probed a human CpG island microarray with DNA from a GC cell line (MKN45) by chromatin immunoprecipitation (ChIP). ChIP-reverse-transcriptase quantitative polymerase chain reaction PCR (RT-qPCR) was used to validate the microarray results. Additionally, mRNA expression levels and the DNA methylation of potential target genes were evaluated by RT-qPCR and methylation-specific PCR (MSP). The moults showed that 134 genes exhibited the highest signal-to-noise ratio of H3-K9 trimethylation over acetylation and 46 genes exhibited the highest signal-to-noise ratio of H3-K9 trimethylation over H3-K4 trimethylation in MKN45 cells. The ChIP-qPCR results agreed with those obtained from the ChIP-chip analysis. Aberrant DNA methylation status and mRNA expression levels were also identified for selected genes (PSD, SMARCC1 and Vps37A) in the GC cell lines. The results suggest that CpG island microarray coupled with ChIP (ChIP-chip) can identify novel targets of gene silencing in GC. Additionally, ChIP-chip is the best approach for assessing the genome-wide status of epigenetic regulation, which may allow for a broader genomic understanding compared to the knowledge that has been accumulated from single-gene studies. PMID:25738530

  16. Genome-wide analysis of histone modifications by ChIP-chip to identify silenced genes in gastric cancer.

    PubMed

    Zhu, Xinjiang; Liu, Jian; Xu, Xiaoyang; Zhang, Chundong; Dai, Dongqiu

    2015-05-01

    The present study aimed to identify novel histone modification markers in gastric cancer (GC) by chromatin immunoprecipitation microarray (ChIP-chip) analysis and to determine whether these markers were able to discriminate between normal and GC cells. We also tested for correlations with DNA methylation. We probed a human CpG island microarray with DNA from a GC cell line (MKN45) by chromatin immunoprecipitation (ChIP). ChIP-reverse-transcriptase quantitative polymerase chain reaction PCR (RT-qPCR) was used to validate the microarray results. Additionally, mRNA expression levels and the DNA methylation of potential target genes were evaluated by RT-qPCR and methylation-specific PCR (MSP). The moults showed that 134 genes exhibited the highest signal-to-noise ratio of H3-K9 trimethylation over acetylation and 46 genes exhibited the highest signal-to-noise ratio of H3-K9 trimethylation over H3-K4 trimethylation in MKN45 cells. The ChIP-qPCR results agreed with those obtained from the ChIP-chip analysis. Aberrant DNA methylation status and mRNA expression levels were also identified for selected genes (PSD, SMARCC1 and Vps37A) in the GC cell lines. The results suggest that CpG island microarray coupled with ChIP (ChIP-chip) can identify novel targets of gene silencing in GC. Additionally, ChIP-chip is the best approach for assessing the genome-wide status of epigenetic regulation, which may allow for a broader genomic understanding compared to the knowledge that has been accumulated from single-gene studies.

  17. Computational method and system for modeling, analyzing, and optimizing DNA amplification and synthesis

    DOEpatents

    Vandersall, Jennifer A.; Gardner, Shea N.; Clague, David S.

    2010-05-04

    A computational method and computer-based system of modeling DNA synthesis for the design and interpretation of PCR amplification, parallel DNA synthesis, and microarray chip analysis. The method and system include modules that address the bioinformatics, kinetics, and thermodynamics of DNA amplification and synthesis. Specifically, the steps of DNA selection, as well as the kinetics and thermodynamics of DNA hybridization and extensions, are addressed, which enable the optimization of the processing and the prediction of the products as a function of DNA sequence, mixing protocol, time, temperature and concentration of species.

  18. Analytical Validation of AmpliChip p53 Research Test for Archival Human Ovarian FFPE Sections.

    PubMed

    Marton, Matthew J; McNamara, Andrew R; Nikoloff, D Michele; Nakao, Aki; Cheng, Jonathan

    2015-01-01

    The p53 tumor suppressor gene (TP53) is reported to be mutated in nearly half of all tumors and plays a central role in genome integrity. Detection of mutations in p53 can be accomplished by many assays, including the AmpliChip p53 Research Test. The AmpliChip p53 Research Test has been successfully used to determine p53 status in hematologic malignancies and fresh frozen solid tissues but there are few reports of using the assay with formalin fixed, paraffin-embedded (FFPE) tissue. The objective of this study was to describe analytical performance characterization of the AmpliChip p53 Research Test to detect p53 mutations in genomic DNA isolated from archival FFPE human ovarian tumor tissues. Method correlation with sequencing showed 96% mutation-wise agreement and 99% chip-wise agreement. We furthermore observed 100% agreement (113/113) of the most prevalent TP53 mutations. Workflow reproducibility was 96.8% across 8 samples, with 2 operators, 2 reagent lots and 2 instruments. Section-to-section reproducibility was 100% for each sample across a 60 μm region of the FFPE block from ovarian tumors. These data indicate that the AmpliChip p53 Research Test is an accurate and reproducible method for detecting mutations in TP53 from archival FFPE human ovarian specimens.

  19. On-chip Extraction of Intracellular Molecules in White Blood Cells from Whole Blood.

    PubMed

    Choi, Jongchan; Hyun, Ji-chul; Yang, Sung

    2015-10-14

    The extraction of virological markers in white blood cells (WBCs) from whole blood--without reagents, electricity, or instruments--is the most important first step for diagnostic testing of infectious diseases in resource-limited settings. Here we develop an integrated microfluidic chip that continuously separates WBCs from whole blood and mechanically ruptures them to extract intracellular proteins and nucleic acids for diagnostic purposes. The integrated chip is assembled with a device that separates WBCs by using differences in blood cell size and a mechanical cell lysis chip with ultra-sharp nanoblade arrays. We demonstrate the performance of the integrated device by quantitatively analyzing the levels of extracted intracellular proteins and genomic DNAs. Our results show that compared with a conventional method, the device yields 120% higher level of total protein amount and similar levels of gDNA (90.3%). To demonstrate its clinical application to human immunodeficiency virus (HIV) diagnostics, the developed chip was used to process blood samples containing HIV-infected cells. Based on PCR results, we demonstrate that the chip can extract HIV proviral DNAs from infected cells with a population as low as 10(2)/μl. These findings suggest that the developed device has potential application in point-of-care testing for infectious diseases in developing countries.

  20. On-chip Extraction of Intracellular Molecules in White Blood Cells from Whole Blood

    PubMed Central

    Choi, Jongchan; Hyun, Ji-chul; Yang, Sung

    2015-01-01

    The extraction of virological markers in white blood cells (WBCs) from whole blood—without reagents, electricity, or instruments—is the most important first step for diagnostic testing of infectious diseases in resource-limited settings. Here we develop an integrated microfluidic chip that continuously separates WBCs from whole blood and mechanically ruptures them to extract intracellular proteins and nucleic acids for diagnostic purposes. The integrated chip is assembled with a device that separates WBCs by using differences in blood cell size and a mechanical cell lysis chip with ultra-sharp nanoblade arrays. We demonstrate the performance of the integrated device by quantitatively analyzing the levels of extracted intracellular proteins and genomic DNAs. Our results show that compared with a conventional method, the device yields 120% higher level of total protein amount and similar levels of gDNA (90.3%). To demonstrate its clinical application to human immunodeficiency virus (HIV) diagnostics, the developed chip was used to process blood samples containing HIV-infected cells. Based on PCR results, we demonstrate that the chip can extract HIV proviral DNAs from infected cells with a population as low as 102/μl. These findings suggest that the developed device has potential application in point-of-care testing for infectious diseases in developing countries. PMID:26464211

  1. On-chip Extraction of Intracellular Molecules in White Blood Cells from Whole Blood

    NASA Astrophysics Data System (ADS)

    Choi, Jongchan; Hyun, Ji-Chul; Yang, Sung

    2015-10-01

    The extraction of virological markers in white blood cells (WBCs) from whole blood—without reagents, electricity, or instruments—is the most important first step for diagnostic testing of infectious diseases in resource-limited settings. Here we develop an integrated microfluidic chip that continuously separates WBCs from whole blood and mechanically ruptures them to extract intracellular proteins and nucleic acids for diagnostic purposes. The integrated chip is assembled with a device that separates WBCs by using differences in blood cell size and a mechanical cell lysis chip with ultra-sharp nanoblade arrays. We demonstrate the performance of the integrated device by quantitatively analyzing the levels of extracted intracellular proteins and genomic DNAs. Our results show that compared with a conventional method, the device yields 120% higher level of total protein amount and similar levels of gDNA (90.3%). To demonstrate its clinical application to human immunodeficiency virus (HIV) diagnostics, the developed chip was used to process blood samples containing HIV-infected cells. Based on PCR results, we demonstrate that the chip can extract HIV proviral DNAs from infected cells with a population as low as 102/μl. These findings suggest that the developed device has potential application in point-of-care testing for infectious diseases in developing countries.

  2. Analytical Validation of AmpliChip p53 Research Test for Archival Human Ovarian FFPE Sections.

    PubMed

    Marton, Matthew J; McNamara, Andrew R; Nikoloff, D Michele; Nakao, Aki; Cheng, Jonathan

    2015-01-01

    The p53 tumor suppressor gene (TP53) is reported to be mutated in nearly half of all tumors and plays a central role in genome integrity. Detection of mutations in p53 can be accomplished by many assays, including the AmpliChip p53 Research Test. The AmpliChip p53 Research Test has been successfully used to determine p53 status in hematologic malignancies and fresh frozen solid tissues but there are few reports of using the assay with formalin fixed, paraffin-embedded (FFPE) tissue. The objective of this study was to describe analytical performance characterization of the AmpliChip p53 Research Test to detect p53 mutations in genomic DNA isolated from archival FFPE human ovarian tumor tissues. Method correlation with sequencing showed 96% mutation-wise agreement and 99% chip-wise agreement. We furthermore observed 100% agreement (113/113) of the most prevalent TP53 mutations. Workflow reproducibility was 96.8% across 8 samples, with 2 operators, 2 reagent lots and 2 instruments. Section-to-section reproducibility was 100% for each sample across a 60 μm region of the FFPE block from ovarian tumors. These data indicate that the AmpliChip p53 Research Test is an accurate and reproducible method for detecting mutations in TP53 from archival FFPE human ovarian specimens. PMID:26125596

  3. Starr: Simple Tiling ARRay analysis of Affymetrix ChIP-chip data

    PubMed Central

    2010-01-01

    Background Chromatin immunoprecipitation combined with DNA microarrays (ChIP-chip) is an assay used for investigating DNA-protein-binding or post-translational chromatin/histone modifications. As with all high-throughput technologies, it requires thorough bioinformatic processing of the data for which there is no standard yet. The primary goal is to reliably identify and localize genomic regions that bind a specific protein. Further investigation compares binding profiles of functionally related proteins, or binding profiles of the same proteins in different genetic backgrounds or experimental conditions. Ultimately, the goal is to gain a mechanistic understanding of the effects of DNA binding events on gene expression. Results We present a free, open-source R/Bioconductor package Starr that facilitates comparative analysis of ChIP-chip data across experiments and across different microarray platforms. The package provides functions for data import, quality assessment, data visualization and exploration. Starr includes high-level analysis tools such as the alignment of ChIP signals along annotated features, correlation analysis of ChIP signals with complementary genomic data, peak-finding and comparative display of multiple clusters of binding profiles. It uses standard Bioconductor classes for maximum compatibility with other software. Moreover, Starr automatically updates microarray probe annotation files by a highly efficient remapping of microarray probe sequences to an arbitrary genome. Conclusion Starr is an R package that covers the complete ChIP-chip workflow from data processing to binding pattern detection. It focuses on the high-level data analysis, e.g., it provides methods for the integration and combined statistical analysis of binding profiles and complementary functional genomics data. Starr enables systematic assessment of binding behaviour for groups of genes that are alingned along arbitrary genomic features. PMID:20398407

  4. An integrated lab-on-chip for rapid identification and simultaneous differentiation of tropical pathogens.

    PubMed

    Tan, Jeslin J L; Capozzoli, Monica; Sato, Mitsuharu; Watthanaworawit, Wanitda; Ling, Clare L; Mauduit, Marjorie; Malleret, Benoît; Grüner, Anne-Charlotte; Tan, Rosemary; Nosten, François H; Snounou, Georges; Rénia, Laurent; Ng, Lisa F P

    2014-01-01

    Tropical pathogens often cause febrile illnesses in humans and are responsible for considerable morbidity and mortality. The similarities in clinical symptoms provoked by these pathogens make diagnosis difficult. Thus, early, rapid and accurate diagnosis will be crucial in patient management and in the control of these diseases. In this study, a microfluidic lab-on-chip integrating multiplex molecular amplification and DNA microarray hybridization was developed for simultaneous detection and species differentiation of 26 globally important tropical pathogens. The analytical performance of the lab-on-chip for each pathogen ranged from 102 to 103 DNA or RNA copies. Assay performance was further verified with human whole blood spiked with Plasmodium falciparum and Chikungunya virus that yielded a range of detection from 200 to 4×105 parasites, and from 250 to 4×107 PFU respectively. This lab-on-chip was subsequently assessed and evaluated using 170 retrospective patient specimens in Singapore and Thailand. The lab-on-chip had a detection sensitivity of 83.1% and a specificity of 100% for P. falciparum; a sensitivity of 91.3% and a specificity of 99.3% for P. vivax; a positive 90.0% agreement and a specificity of 100% for Chikungunya virus; and a positive 85.0% agreement and a specificity of 100% for Dengue virus serotype 3 with reference methods conducted on the samples. Results suggested the practicality of an amplification microarray-based approach in a field setting for high-throughput detection and identification of tropical pathogens.

  5. Infrared vertically-illuminated photodiode for chip alignment feedback

    NASA Astrophysics Data System (ADS)

    Alloatti, L.; Ram, R. J.

    2016-08-01

    We report on vertically-illuminated photodiodes fabricated in the GlobalFoundries 45nm 12SOI node and on a packaging concept for optically-interconnected chips. The photodiodes are responsive at 1180 nm -a wavelength currently used in chip-to-chip communications. They have further a wide field-of-view which enables chip-to-board positional feedback in chip-board assemblies. Monolithic integration enables on-chip processing of the positional data.

  6. Wax-bonding 3D microfluidic chips.

    PubMed

    Gong, Xiuqing; Yi, Xin; Xiao, Kang; Li, Shunbo; Kodzius, Rimantas; Qin, Jianhua; Wen, Weijia

    2010-10-01

    We report a simple, low-cost and detachable microfluidic chip incorporating easily accessible paper, glass slides or other polymer films as the chip materials along with adhesive wax as the recycling bonding material. We use a laser to cut through the paper or film to form patterns and then sandwich the paper and film between glass sheets or polymer membranes. The hot-melt adhesive wax can realize bridge bonding between various materials, for example, paper, polymethylmethacrylate (PMMA) film, glass sheets, or metal plate. The bonding process is reversible and the wax is reusable through a melting and cooling process. With this process, a three-dimensional (3D) microfluidic chip is achievable by vacuating and venting the chip in a hot-water bath. To study the biocompatibility and applicability of the wax-based microfluidic chip, we tested the PCR compatibility with the chip materials first. Then we applied the wax-paper based microfluidic chip to HeLa cell electroporation (EP). Subsequently, a prototype of a 5-layer 3D chip was fabricated by multilayer wax bonding. To check the sealing ability and the durability of the chip, green fluorescence protein (GFP) recombinant Escherichia coli (E. coli) bacteria were cultured, with which the chemotaxis of E. coli was studied in order to determine the influence of antibiotic ciprofloxacin concentration on the E. coli migration.

  7. Fermilab silicon strip readout chip for BTev

    SciTech Connect

    Yarema, Raymond; Hoff, Jim; Mekkaoui, Abderrezak; Manghisoni, Massimo; Re, Valerio; Angeleri, Valentina; Manfredi, Pier Francesco; Ratti, Lodovico; Speziali, Valeria; /Fermilab /Bergamo U. /INFN, Pavia /Pavia U.

    2005-05-01

    A chip has been developed for reading out the silicon strip detectors in the new BTeV colliding beam experiment at Fermilab. The chip has been designed in a 0.25 {micro}m CMOS technology for high radiation tolerance. Numerous programmable features have been added to the chip, such as setup for operation at different beam crossing intervals. A full size chip has been fabricated and successfully tested. The design philosophy, circuit features, and test results are presented in this paper.

  8. Bulk-micromachined submicroliter-volume PCR chip with very rapid thermal response and low power consumption.

    PubMed

    Lee, Dae-Sik; Park, Se Ho; Yang, Haesik; Chung, Kwang-Hyo; Yoon, Tae Hwan; Kim, Sung-Jin; Kim, Kyuwon; Kim, Youn Tae

    2004-08-01

    The current paper describes the design, fabrication, and testing of a micromachined submicroliter-volume polymerase chain reaction (PCR) chip with a fast thermal response and very low power consumption. The chip consists of a bulk-micromachined Si component and hot-embossed poly(methyl methacrylate)(PMMA) component. The Si component contains an integral microheater and temperature sensor on a thermally well-isolated membrane, while the PMMA component contains a submicroliter-volume PCR chamber, valves, and channels. The micro hot membrane under the submicroliter-volume chamber is a silicon oxide/silicon nitride/silicon oxide (O/N/O) diaphragm with a thickness of 1.9 microm, resulting in a very low thermal mass. In experiments, the proposed chip only required 45 mW to heat the reaction chamber to 92 degrees C, the denaturation temperature of DNA, plus the heating and cooling rates are about 80 degrees C s(-1) and 60 degrees C s(-1), respectively. We validated, from the fluorescence results from DNA stained with SYBR Green I, that the proposed chip amplified the DNA from vector clone, containing tumor suppressor gene BRCA 1 (127 base pairs at 11th exon), after 30 thermal cycles of 3 s, 5 s, and 5 s at 92 degrees C, 55 degrees C, and 72 degrees C, respectively, in a 200 nL-volume chamber. As for specificity of DNA products, owing to difficulty in analyzing the very small volume PCR results from the micro chip, we vicariously employed the larger volume PCR products after cycling with the same sustaining temperatures as with the micro chip but with much slower ramping rates (3.3 degrees C s(-1) when rising, 2.5 degrees C s(-1) when cooling) within circa 20 minutes on a commercial PCR machine and confirmed the specificity to BRCA 1 (127 base pairs) with agarose gel electrophoresis. Accordingly, the fabricated micro chip demonstrated a very low power consumption and rapid thermal response, both of which are crucial to the development of a fully integrated and battery

  9. Optofluidic chips with nanochannels for dynamic molecular detection using enhanced fluorescence

    PubMed Central

    Postigo, P. A.; Alvaro, R.; Juarros, A.; Merino, S.

    2016-01-01

    The fabrication of a novel optofluidic chip using nanochannels optimized for DNA-stretched molecules and optical detection by enhanced fluorescence is reported. The chips are composed of a series of microchannels that allow the transport of molecules in the femto-liter per second inside a fluid or gas. The nanochannels are surrounded by a photonic crystal structure to enhance the emission of fluorescent light from the molecules, which can travel along the nanochannel, allowing for enhanced optical detection of the molecules in motion. The photonic crystal structure provides an enhancement up to 2.5 times in the light emitted from fluorescent molecules inside the nanochannels which increases to around 250 when normalized to the area of the nanochannels emitting fluorescence. The results may help to the detection of fluorescent molecules (like marked-DNA) in series by speeding it and allowing the use of less sophisticated equipment. PMID:27699099

  10. Optofluidic chips with nanochannels for dynamic molecular detection using enhanced fluorescence

    PubMed Central

    Postigo, P. A.; Alvaro, R.; Juarros, A.; Merino, S.

    2016-01-01

    The fabrication of a novel optofluidic chip using nanochannels optimized for DNA-stretched molecules and optical detection by enhanced fluorescence is reported. The chips are composed of a series of microchannels that allow the transport of molecules in the femto-liter per second inside a fluid or gas. The nanochannels are surrounded by a photonic crystal structure to enhance the emission of fluorescent light from the molecules, which can travel along the nanochannel, allowing for enhanced optical detection of the molecules in motion. The photonic crystal structure provides an enhancement up to 2.5 times in the light emitted from fluorescent molecules inside the nanochannels which increases to around 250 when normalized to the area of the nanochannels emitting fluorescence. The results may help to the detection of fluorescent molecules (like marked-DNA) in series by speeding it and allowing the use of less sophisticated equipment.

  11. On chip shapeable optical tweezers

    NASA Astrophysics Data System (ADS)

    Renaut, C.; Cluzel, B.; Dellinger, J.; Lalouat, L.; Picard, E.; Peyrade, D.; Hadji, E.; de Fornel, F.

    2013-07-01

    Particles manipulation with optical forces is known as optical tweezing. While tweezing in free space with laser beams was established in the 1980s, integrating the optical tweezers on a chip is a challenging task. Recent experiments with plasmonic nanoantennas, microring resonators, and photonic crystal nanocavities have demonstrated optical trapping. However, the optical field of a tweezer made of a single microscopic resonator cannot be shaped. So far, this prevents from optically driven micromanipulations. Here we propose an alternative approach where the shape of the optical trap can be tuned by the wavelength in coupled nanobeam cavities. Using these shapeable tweezers, we present micromanipulation of polystyrene microspheres trapped on a silicon chip. These results show that coupled nanobeam cavities are versatile building blocks for optical near-field engineering. They open the way to much complex integrated tweezers using networks of coupled nanobeam cavities for particles or bio-objects manipulation at a larger scale.

  12. On-chip plasmonic spectrometer.

    PubMed

    Tsur, Yuval; Arie, Ady

    2016-08-01

    We report a numerical and experimental study of an on-chip optical spectrometer, utilizing propagating surface plasmon polaritons in the telecom spectral range. The device is based on two holographic gratings, one for coupling, and the other for decoupling free-space radiation with the surface plasmons. This 800 μm×100 μm on-chip spectrometer resolves 17 channels spectrally separated by 3.1 nm, spanning a freely tunable spectral window, and is based on standard lithography fabrication technology. We propose two potential applications for this new device; the first employs the holographic control over the amplitude and phase of the input spectrum, for intrinsically filtering unwanted frequencies, like pump radiation in Raman spectroscopy. The second prospect utilizes the unique plasmonic field enhancement at the metal-dielectric boundary for the spectral analysis of very small samples (e.g., Mie scatterers) placed between the two gratings.

  13. On-chip plasmonic spectrometer.

    PubMed

    Tsur, Yuval; Arie, Ady

    2016-08-01

    We report a numerical and experimental study of an on-chip optical spectrometer, utilizing propagating surface plasmon polaritons in the telecom spectral range. The device is based on two holographic gratings, one for coupling, and the other for decoupling free-space radiation with the surface plasmons. This 800 μm×100 μm on-chip spectrometer resolves 17 channels spectrally separated by 3.1 nm, spanning a freely tunable spectral window, and is based on standard lithography fabrication technology. We propose two potential applications for this new device; the first employs the holographic control over the amplitude and phase of the input spectrum, for intrinsically filtering unwanted frequencies, like pump radiation in Raman spectroscopy. The second prospect utilizes the unique plasmonic field enhancement at the metal-dielectric boundary for the spectral analysis of very small samples (e.g., Mie scatterers) placed between the two gratings. PMID:27472609

  14. Tetrameric assembly of CHIP28 water channels in liposomes and cell membranes: a freeze-fracture study.

    PubMed

    Verbavatz, J M; Brown, D; Sabolić, I; Valenti, G; Ausiello, D A; Van Hoek, A N; Ma, T; Verkman, A S

    1993-11-01

    Channel forming integral protein of 28 kD (CHIP28) functions as a water channel in erythrocytes, kidney proximal tubule and thin descending limb of Henle. CHIP28 morphology was examined by freeze-fracture EM in proteoliposomes reconstituted with purified CHIP28, CHO cells stably transfected with CHIP28k cDNA, and rat kidney tubules. Liposomes reconstituted with HPLC-purified CHIP28 from human erythrocytes had a high osmotic water permeability (Pf0.04 cm/s) that was inhibited by HgCl2. Freeze-fracture replicas showed a fairly uniform set of intramembrane particles (IMPs); no IMPs were observed in liposomes without incorporated protein. By rotary shadowing, the IMPs had a diameter of 8.5 +/- 1.3 nm (mean +/- SD); many IMPs consisted of a distinct arrangement of four smaller subunits surrounding a central depression. IMPs of similar size and appearance were seen on the P-face of plasma membranes from CHIP28k-transfected (but not mock-transfected) CHO cells, rat thin descending limb (TDL) of Henle, and S3 segment of proximal straight tubules. A distinctive network of complementary IMP imprints was observed on the E-face of CHIP28-containing plasma membranes. The densities of IMPs in the size range of CHIP28 IMPs, determined by non-linear regression, were (in IMPs/microns 2): 2,494 in CHO cells, 5,785 in TDL, and 1,928 in proximal straight tubules; predicted Pf, based on the CHIP28 single channel water permeability of 3.6 x 10(-14) cm3/S (10 degrees C), was in good agreement with measured Pf of 0.027 cm/S, 0.075 cm/S, and 0.031 cm/S, respectively, in these cell types. Assuming that each CHIP28 monomer is a right cylindrical pore of length 5 nm and density 1.3 g/cm3, the monomer diameter would be 3.2 nm; a symmetrical arrangement of four cylinders would have a greatest diameter of 7.2 nm, which after correction for the thickness of platinum deposit, is similar to the measured IMP diameter of approximately 8.5 nm. These results provide a morphological signature for CHIP28

  15. Identification of polymorphic and off-target probe binding sites on the Illumina Infinium MethylationEPIC BeadChip.

    PubMed

    McCartney, Daniel L; Walker, Rosie M; Morris, Stewart W; McIntosh, Andrew M; Porteous, David J; Evans, Kathryn L

    2016-09-01

    Genome-wide analysis of DNA methylation has now become a relatively inexpensive technique thanks to array-based methylation profiling technologies. The recently developed Illumina Infinium MethylationEPIC BeadChip interrogates methylation at over 850,000 sites across the human genome, covering 99% of RefSeq genes. This array supersedes the widely used Infinium HumanMethylation450 BeadChip, which has permitted insights into the relationship between DNA methylation and a wide range of conditions and traits. Previous research has identified issues with certain probes on both the HumanMethylation450 BeadChip and its predecessor, the Infinium HumanMethylation27 BeadChip, which were predicted to affect array performance. These issues concerned probe-binding specificity and the presence of polymorphisms at target sites. Using in silico methods, we have identified probes on the Infinium MethylationEPIC BeadChip that are predicted to (i) measure methylation at polymorphic sites and (ii) hybridise to multiple genomic regions. We intend these resources to be used for quality control procedures when analysing data derived from this platform. PMID:27330998

  16. DNA nanomachines.

    PubMed

    Bath, Jonathan; Turberfield, Andrew J

    2007-05-01

    We are learning to build synthetic molecular machinery from DNA. This research is inspired by biological systems in which individual molecules act, singly and in concert, as specialized machines: our ambition is to create new technologies to perform tasks that are currently beyond our reach. DNA nanomachines are made by self-assembly, using techniques that rely on the sequence-specific interactions that bind complementary oligonucleotides together in a double helix. They can be activated by interactions with specific signalling molecules or by changes in their environment. Devices that change state in response to an external trigger might be used for molecular sensing, intelligent drug delivery or programmable chemical synthesis. Biological molecular motors that carry cargoes within cells have inspired the construction of rudimentary DNA walkers that run along self-assembled tracks. It has even proved possible to create DNA motors that move autonomously, obtaining energy by catalysing the reaction of DNA or RNA fuels.

  17. Technical Considerations in using DNA Microarrays to Define Regulons

    PubMed Central

    Rhodius, Virgil A.; Wade, Joseph T.

    2009-01-01

    Transcription is the major regulatory target of gene expression in bacteria, and is controlled by many regulatory proteins and RNAs. Microarrays are a powerful tool to study the regulation of transcription on a genomic scale. Here we describe the use of transcription profiling and ChIP-chip to study transcriptional regulation in bacteria. Transcription profiling determines the outcome of regulatory events whereas ChIP-chip identifies the protein-DNA interactions that determine these events. Together they can provide detailed information on transcriptional regulatory systems. PMID:18955146

  18. Methylsorb: a simple method for quantifying DNA methylation using DNA-gold affinity interactions.

    PubMed

    Sina, Abu Ali Ibn; Carrascosa, Laura G; Palanisamy, Ramkumar; Rauf, Sakandar; Shiddiky, Muhammad J A; Trau, Matt

    2014-10-21

    The analysis of DNA methylation is becoming increasingly important both in the clinic and also as a research tool to unravel key epigenetic molecular mechanisms in biology. Current methodologies for the quantification of regional DNA methylation (i.e., the average methylation over a region of DNA in the genome) are largely affected by comprehensive DNA sequencing methodologies which tend to be expensive, tedious, and time-consuming for many applications. Herein, we report an alternative DNA methylation detection method referred to as "Methylsorb", which is based on the inherent affinity of DNA bases to the gold surface (i.e., the trend of the affinity interactions is adenine > cytosine ≥ guanine > thymine).1 Since the degree of gold-DNA affinity interaction is highly sequence dependent, it provides a new capability to detect DNA methylation by simply monitoring the relative adsorption of bisulfite treated DNA sequences onto a gold chip. Because the selective physical adsorption of DNA fragments to gold enable a direct read-out of regional DNA methylation, the current requirement for DNA sequencing is obviated. To demonstrate the utility of this method, we present data on the regional methylation status of two CpG clusters located in the EN1 and MIR200B genes in MCF7 and MDA-MB-231 cells. The methylation status of these regions was obtained from the change in relative mass on gold surface with respect to relative adsorption of an unmethylated DNA source and this was detected using surface plasmon resonance (SPR) in a label-free and real-time manner. We anticipate that the simplicity of this method, combined with the high level of accuracy for identifying the methylation status of cytosines in DNA, could find broad application in biology and diagnostics.

  19. Multiparameter Lab-on-a-Chip flow cytometry of the cell cycle.

    PubMed

    Skommer, Joanna; Akagi, Jin; Takeda, Kazuo; Fujimura, Yuu; Khoshmanesh, Khashayar; Wlodkowic, Donald

    2013-04-15

    Multiparameter analysis of apoptosis in relation to cell cycle position is helpful in exploring mechanism of action of anticancer drugs that target specific molecular cogs of the cell cycle. This work demonstrates a new rationale for using microfluidic Lab-on-a-Chip flow cytometry (μFCM) with a simple 2D hydrodynamic focusing for the multiparameter analysis of apoptosis and DNA ploidy analysis in human hematopoietic cancer cells. The microfluidic system employs disposable microfluidic cartridges fabricated using injection moulding in optically transparent poly(methylmethacrylate). The dedicated and miniaturized electronic hardware interface enables up to six parameter detections using a combination of spatially separated solid-state 473 nm (10 mW) and 640 nm (20 mW) lasers and x-y stage for rapid laser alignment adjustment. We provide evidence that the simple 2D flow focusing on a chip-based device is sufficient to measure cellular DNA content in both fixed and living tumor cells. The feasibility of using the μFCM system for multiparameter analysis of caspase activation and dissipation of mitochondrial inner membrane potential (ΔΨ(m) loss) in relation to DNA content is also demonstrated. The data shows that straightforward microfluidic chip designs are sufficient to acquire high quality biological data when combined with sophisticated electronic interfaces. They can be a viable alternative to conventional FCM for multiparameter detection of programmed cell death.

  20. Sensitive determination of DNA based on the interaction between prulifloxacin-terbium(III) complex and DNA.

    PubMed

    Wu, Ting; Fang, Biyun; Chang, Lin; Liu, Min; Chen, Fang

    2013-01-01

    A simple spectrofluorimetric method is described for the determination of DNA, based on its enhancement of the fluorescence intensity of prulifloxacin (PUFX)-Tb(3+). The luminescence intensity of the PUFX-Tb(3+) complex increased up to 10-fold after adding DNA. The excitation and emission wavelengths were 345 and 545 nm, respectively. Under optimum conditions, variations in the fluorescence intensity showed a good linear relationship with the concentration of hsDNA in the range of 3.0 × 10(-9) to 1.0 × 10(-6) g/mL, with a correlation coefficient (R) of 0.997, and the detection limit was 2.1 × 10(-9) g/mL. The method was successfully applied to the determination of DNA in synthetic samples, and recoveries were in the range 97.3-102.0%. The mechanism of fluorescence enhancement of the PUFX-Tb(3+) complex by DNA is also discussed. The mechanism may involve formation of a ternary complex mainly by intercalation binding together with weak electrostatic interaction, which will increase the energy transition from ligand to Tb(3+), increasing the rigidity of the complex, and decreasing the radiationless energy loss through O-H vibration of the H2O molecule in the PUFX-Tb(3+) complex. Compared with the previous DNA probes, the proposed method is not only more robust and friendly to the environment, but also of relatively higher sensitivity.

  1. CHIP has a protective role against oxidative stress-induced cell death through specific regulation of Endonuclease G

    PubMed Central

    Lee, J S; Seo, T W; Yi, J H; Shin, K S; Yoo, S J

    2013-01-01

    Oxidative stress is implicated in carcinogenesis, aging, and neurodegenerative diseases. The E3 ligase C terminus of Hsc-70 interacting protein (CHIP) has a protective role against various stresses by targeting damaged proteins for proteasomal degradation, and thus maintains protein quality control. However, the detailed mechanism by which CHIP protects cells from oxidative stress has not been demonstrated. Here, we show that depletion of CHIP led to elevated Endonuclease G (EndoG) levels and enhanced cell death upon oxidative stress. In contrast, CHIP overexpression reduced EndoG levels, and resulted in reduced or no oxidative stress-induced cell death in cancer cells and primary rat cortical neurons. Under normal conditions Hsp70 mediated the interaction between EndoG and CHIP, downregulating EndoG levels in a Hsp70/proteasome-dependent manner. However, under oxidative stress Hsp70 no longer interacted with EndoG, and the stabilized EndoG translocated to the nucleus and degraded chromosomal DNA. Our data suggest that regulation of the level of EndoG by CHIP in normal conditions may determine the sensitivity to cell death upon oxidative stress. Indeed, injection of H2O2 into the rat brain markedly increased cell death in aged mice compared with young mice, which correlated with elevated levels of EndoG and concurrent downregulation of CHIP in aged mice. Taken together, our findings demonstrate a novel protective mechanism of CHIP against oxidative stress through regulation of EndoG, and provide an opportunity to modulate oxidative stress-induced cell death in cancer and aging. PMID:23764847

  2. HPV Direct Flow CHIP: a new human papillomavirus genotyping method based on direct PCR from crude-cell extracts.

    PubMed

    Herraez-Hernandez, Elsa; Alvarez-Perez, Martina; Navarro-Bustos, Gloria; Esquivias, Javier; Alonso, Sonia; Aneiros-Fernandez, Jose; Lacruz-Pelea, Cesar; Sanchez-Aguera, Magdalena; Santamaria, Javier Saenz; de Antonio, Jesus Chacon; Rodriguez-Peralto, Jose Luis

    2013-10-01

    HPV Direct Flow CHIP is a newly developed test for identifying 18 high-risk and 18 low-risk human papillomavirus (HPV) genotypes. It is based on direct PCR from crude-cell extracts, automatic flow-through hybridization, and colorimetric detection. The aim of this study was to evaluate the performance of HPV Direct Flow CHIP in the analysis of 947 samples from routine cervical screening or the follow-up of abnormal Pap smears. The specimens were dry swab samples, liquid-based cytology samples, or formalin-fixed paraffin-embedded tissues. The genotype distribution was in agreement with known epidemiological data for the Spanish population. Three different subgroups of the samples were also tested by Linear Array (LA) HPV Genotyping Test (n=108), CLART HPV2 (n=82), or Digene Hybrid Capture 2 (HC2) HPV DNA Test (n=101). HPV positivity was 73.6% by HPV Direct Flow CHIP versus 67% by LA, 65.9% by HPV Direct Flow CHIP versus 59.8% by CLART, and 62.4% by HPV Direct Flow CHIP versus 42.6% by HC2. HPV Direct Flow CHIP showed a positive agreement of 88.6% with LA (k=0.798), 87.3% with CLART (k=0.818), and 68.2% with HC2 (k=0.618). In conclusion, HPV Direct Flow CHIP results were comparable with those of the other methods tested. Although further investigation is needed to compare the performance of this new test with a gold-standard reference method, these preliminary findings evidence the potential value of HPV Direct Flow CHIP in HPV vaccinology and epidemiology studies.

  3. Electrokinetic concentration of DNA polymers in nanofluidic channels.

    PubMed

    Stein, Derek; Deurvorst, Zeno; van der Heyden, Frank H J; Koopmans, Wiepke J A; Gabel, Alan; Dekker, Cees

    2010-03-10

    DNA molecules can be concentrated in a narrow region of a nanochannel when driven electrokinetically in submillimolar salt solutions. Transport experiments and theoretical modeling reveal the interplay of electrophoresis, electro-osmosis, and the unique statistical properties of confined polymers that lead to DNA aggregation. A finite conductance through the bulk of the device also plays a crucial role by influencing the electric fields in the nanochannel. We build on this understanding by demonstrating how a nanofluidic device with integrated electrodes can preconcentrate DNA at selected locations and at physiological salt concentrations that are relevant to lab-on-a-chip applications. PMID:20151696

  4. [DNA computing].

    PubMed

    Błasiak, Janusz; Krasiński, Tadeusz; Popławski, Tomasz; Sakowski, Sebastian

    2011-01-01

    Biocomputers can be an alternative for traditional "silicon-based" computers, which continuous development may be limited due to further miniaturization (imposed by the Heisenberg Uncertainty Principle) and increasing the amount of information between the central processing unit and the main memory (von Neuman bottleneck). The idea of DNA computing came true for the first time in 1994, when Adleman solved the Hamiltonian Path Problem using short DNA oligomers and DNA ligase. In the early 2000s a series of biocomputer models was presented with a seminal work of Shapiro and his colleguas who presented molecular 2 state finite automaton, in which the restriction enzyme, FokI, constituted hardware and short DNA oligomers were software as well as input/output signals. DNA molecules provided also energy for this machine. DNA computing can be exploited in many applications, from study on the gene expression pattern to diagnosis and therapy of cancer. The idea of DNA computing is still in progress in research both in vitro and in vivo and at least promising results of these research allow to have a hope for a breakthrough in the computer science. PMID:21735816

  5. Microchannel cooling of face down bonded chips

    DOEpatents

    Bernhardt, A.F.

    1993-06-08

    Microchannel cooling is applied to flip-chip bonded integrated circuits, in a manner which maintains the advantages of flip-chip bonds, while overcoming the difficulties encountered in cooling the chips. The technique is suited to either multi chip integrated circuit boards in a plane, or to stacks of circuit boards in a three dimensional interconnect structure. Integrated circuit chips are mounted on a circuit board using flip-chip or control collapse bonds. A microchannel structure is essentially permanently coupled with the back of the chip. A coolant delivery manifold delivers coolant to the microchannel structure, and a seal consisting of a compressible elastomer is provided between the coolant delivery manifold and the microchannel structure. The integrated circuit chip and microchannel structure are connected together to form a replaceable integrated circuit module which can be easily decoupled from the coolant delivery manifold and the circuit board. The coolant supply manifolds may be disposed between the circuit boards in a stack and coupled to supplies of coolant through a side of the stack.

  6. Microluminometer chip and method to measure bioluminescence

    SciTech Connect

    Simpson, Michael L; Paulus, Michael J; Sayler, Gary S; Applegate, Bruce M; Ripp, Steven A

    2008-05-13

    An integrated microluminometer includes an integrated circuit chip having at least one n-well/p-substrate junction photodetector for converting light received into a photocurrent, and a detector on the chip for processing the photocurrent. A distributed electrode configuration including a plurality of spaced apart electrodes disposed on an active region of the photodetector is preferably used to raise efficiency.

  7. Radiation Behavior of Analog Neural Network Chip

    NASA Technical Reports Server (NTRS)

    Langenbacher, H.; Zee, F.; Daud, T.; Thakoor, A.

    1996-01-01

    A neural network experiment conducted for the Space Technology Research Vehicle (STRV-1) 1-b launched in June 1994. Identical sets of analog feed-forward neural network chips was used to study and compare the effects of space and ground radiation on the chips. Three failure mechanisms are noted.

  8. Teaching Quality Control with Chocolate Chip Cookies

    ERIC Educational Resources Information Center

    Baker, Ardith

    2014-01-01

    Chocolate chip cookies are used to illustrate the importance and effectiveness of control charts in Statistical Process Control. By counting the number of chocolate chips, creating the spreadsheet, calculating the control limits and graphing the control charts, the student becomes actively engaged in the learning process. In addition, examining…

  9. Multimedia-Based Chip Design Education.

    ERIC Educational Resources Information Center

    Catalkaya, Tamer; Golze, Ulrich

    This paper focuses on multimedia computer-based training programs on chip design. Their development must be fast and economical, in order to be affordable by technical university institutions. The self-produced teaching program Illusion, which demonstrates a monitor controller as an example of a small but complete chip design, was implemented to…

  10. Lab-on a-Chip

    NASA Technical Reports Server (NTRS)

    2003-01-01

    Helen Cole, the project manager for the Lab-on-a-Chip Applications Development program, and Lisa Monaco, the project scientist for the program, insert a lab on a chip into the Caliper 42 which is specialized equipment that controls processes on commercial chips to support development of lab-on-a-chip applications. The system has special microscopes and imaging systems, so scientists can process and study different types of fluid, chemical, and medical tests conducted on chips. For example, researchers have examined fluorescent bacteria as it flows through the chips' fluid channels or microfluidic capillaries. Researchers at NASA's Marshall Space Flight Center (MSFC) in Huntsville, Alabama, have been studying how the lab-on-a-chip technology can be used for microbial detection, water quality monitoring, and detecting biosignatures of past or present life on Mars. The Marshall Center team is also collaborating with scientists at other NASA centers and at universities to develop custom chip designs for not only space applications, but for many Earth applications, such as for detecting deadly microbes in heating and air systems. (NASA/MSFC/D.Stoffer)

  11. Oligonucleotides direct synthesis on porous silicon chip.

    PubMed

    De Stefano, Luca; De Tommasi, Edoardo; Rea, Ilaria; Rotiroti, Lucia; Giangrande, Luca; Oliviero, Giorgia; Borbone, Nicola; Galeone, Aldo; Piccialli, Gennaro

    2008-01-01

    A solid phase oligonucleotide (ON) synthesis on porous silicon (PSi) chip is presented. The prepared Si-OH surface were analyzed by FT-IR and the OH functions were quantified by reaction with 3'-phosphoramidite nucleotide building block. Short ONs were synthesized on the chip surface and the coupling yields evaluated. PMID:18776583

  12. Electrothermal modeling of silicon PCR chips

    NASA Astrophysics Data System (ADS)

    Cui, Zheng; Zhao, Zhan; Xia, Shanhong

    2001-04-01

    Polymerase chain reaction (PCR) on a microchip has drawn considerable attention in recent years. Although a microchip can have must fast heating and cooling rate, the delicacy in its structure makes the PCR experiment difficult and cracks often occurs particularly for the thin membrane type of PCR chips. Electrothermal modeling of PCR chips is presented using commercial MEMS software tool IntelliSuiteTM, with the aim of identifying the problems encountered in experiment and finding an optimum chip structure. Heating characteristics of four different heater designs have been compared, so have the PCR chambers with fixed frame and with suspended frame. The thermal stress analysis has shown that the structure and heater design can make significant difference in heating characteristics and in reducing the failure of PCR chips. The computer simulation has confirmed what has been found in experiment the reason of membrane cracks. Improvement in PCR chip design has been proposed.

  13. Carbon Nanotube Amperometric Chips with Pneumatic Micropumps

    NASA Astrophysics Data System (ADS)

    Tsujita, Yuichi; Maehashi, Kenzo; Matsumoto, Kazuhiko; Chikae, Miyuki; Torai, Soichiro; Takamura, Yuzuru; Tamiya, Eiichi

    2008-04-01

    We fabricated carbon nanotube (CNT) amperometric chips with pneumatic micropumps by the combination of amperometric biosensors based on CNT-arrayed electrodes and microchannels with pneumatic micropumps made of poly(dimethylsiloxane). On the chip, phosphate buffer solution and potassium ferricyanide, K3[Fe(CN)6], were introduced into the CNT electrodes using each pneumatic micropump and electrochemically measured by differential pulse voltammetry. The results indicate that our chip can automatically exchange reagents on the CNT electrodes and clearly detect molecules. Moreover, by modifying the CNT electrodes with enzyme glucose oxidase, glucose molecules could be detected using our chips by cyclic voltammetry and chronoamperometry. We conclude that microfluidic chips with CNT-arrayed electrodes are a promising candidate for the development of hand-held electrochemical biosensors.

  14. A novel CMOS electric field imager for lab-on-a-chip and biomedical applications

    NASA Astrophysics Data System (ADS)

    Ghallab, Yehya Hassan

    Lab-on-a-chip technology is exciting the interest of scientists in many areas. This technology can be used to synthesize chemicals, biological, cancer cells, and DNA efficiently and economically. Dielectrophoresis (DEP) is a fairly well known phenomenon for manipulating and controlling neutral particles. Recently there have been reports of the merging of the robustness of both lab-on-a-chip and DEP phenomenon, such that lab-on-a-chip based on DEP phenomenon has appeared. In support of this research work, this thesis discusses the development of a novel CMOS electric field imager for lab-on-a-chip applications. The imager contains both the sensing and actuation parts in a single chip. The actuation part is a quadupole electrode to apply the required DEP electric field. The sensing part consists of an array of novel electric field sensors, named Differential Electric Field Sensitive Field Effect Transistor (DeFET). Experimental and simulation results show that the chip can process the sensing and the actuation junctions simultaneously and it can trap, concentrate, and quantify biocells. Moreover, the proposed electric field imager can work with the biological systems at the cell level and it can extract real time information about the biological cells behavior using direct measurements. Finally, a novel Current-mode Instrumentation Amplifier (CMIA) is presented that utilizes an Operational Floating Current Conveyor (OFCC) as a basic building block. The proposed CMIA has been analyzed, simulated and experimentally tested. The experimental results verify that the proposed CMIA outperforms existing CMIAs in terms of the number of basic building blocks used, differential gain and CMRR.

  15. Rapid enrichment of leucocytes and genomic DNA from blood based on bifunctional core shell magnetic nanoparticles

    NASA Astrophysics Data System (ADS)

    Xie, Xin; Nie, Xiaorong; Yu, Bingbin; Zhang, Xu

    2007-04-01

    A series of protocols are proposed to extract genomic DNA from whole blood at different scales using carboxyl-functionalized magnetic nanoparticles as solid-phase absorbents. The enrichment of leucocytes and the adsorption of genomic DNA can be achieved with the same carboxyl-functionalized magnetic nanoparticles. The DNA bound to the bead surfaces can be used directly as PCR templates. By coupling cell separation and DNA purification, the whole operation can be accomplished in a few minutes. Our simplified protocols proved to be rapid, low cost, and biologically and chemically non-hazardous, and are therefore promising for microfabrication of a DNA-preparation chip and routine laboratory use.

  16. Transcriptional regulation by CHIP/LDB complexes.

    PubMed

    Bronstein, Revital; Levkovitz, Liron; Yosef, Nir; Yanku, Michaela; Ruppin, Eytan; Sharan, Roded; Westphal, Heiner; Oliver, Brian; Segal, Daniel

    2010-08-12

    It is increasingly clear that transcription factors play versatile roles in turning genes "on" or "off" depending on cellular context via the various transcription complexes they form. This poses a major challenge in unraveling combinatorial transcription complex codes. Here we use the powerful genetics of Drosophila combined with microarray and bioinformatics analyses to tackle this challenge. The nuclear adaptor CHIP/LDB is a major developmental regulator capable of forming tissue-specific transcription complexes with various types of transcription factors and cofactors, making it a valuable model to study the intricacies of gene regulation. To date only few CHIP/LDB complexes target genes have been identified, and possible tissue-dependent crosstalk between these complexes has not been rigorously explored. SSDP proteins protect CHIP/LDB complexes from proteasome dependent degradation and are rate-limiting cofactors for these complexes. By using mutations in SSDP, we identified 189 down-stream targets of CHIP/LDB and show that these genes are enriched for the binding sites of APTEROUS (AP) and PANNIER (PNR), two well studied transcription factors associated with CHIP/LDB complexes. We performed extensive genetic screens and identified target genes that genetically interact with components of CHIP/LDB complexes in directing the development of the wings (28 genes) and thoracic bristles (23 genes). Moreover, by in vivo RNAi silencing we uncovered novel roles for two of the target genes, xbp1 and Gs-alpha, in early development of these structures. Taken together, our results suggest that loss of SSDP disrupts the normal balance between the CHIP-AP and the CHIP-PNR transcription complexes, resulting in down-regulation of CHIP-AP target genes and the concomitant up-regulation of CHIP-PNR target genes. Understanding the combinatorial nature of transcription complexes as presented here is crucial to the study of transcription regulation of gene batteries required for

  17. Dancing DNA.

    ERIC Educational Resources Information Center

    Pennisi, Elizabeth

    1991-01-01

    An imaging technique that uses fluorescent dyes and allows scientists to track DNA as it moves through gels or in solution is described. The importance, opportunities, and implications of this technique are discussed. (KR)

  18. DNA Dynamics.

    ERIC Educational Resources Information Center

    Warren, Michael D.

    1997-01-01

    Explains a method to enable students to understand DNA and protein synthesis using model-building and role-playing. Acquaints students with the triplet code and transcription. Includes copies of the charts used in this technique. (DDR)

  19. Selective Trapping of DNA Using Glass Microcapillaries.

    PubMed

    Rempfer, Georg; Ehrhardt, Sascha; Laohakunakorn, Nadanai; Davies, Gary B; Keyser, Ulrich F; Holm, Christian; de Graaf, Joost

    2016-08-23

    We show experimentally that an inexpensive glass microcapillary can accumulate λ-phage DNA at its tip and deliver the DNA into the capillary using a combination of electro-osmotic flow, pressure-driven flow, and electrophoresis. We develop an efficient simulation model based on the electrokinetic equations and the finite-element method to explain this phenomenon. As a proof of concept for the generality of this trapping mechanism we use our numerical model to explore the effect of the salt concentration, the capillary surface charge, the applied voltage, the pressure difference, and the mobility of the analyte molecules. Our results indicate that the simple microcapillary system has the potential to capture a wide range of analyte molecules based on their electrophoretic mobility that extends well beyond our experimental example of λ-phage DNA. Our method for separation and preconcentration of analytes therefore has implications for the development of low-cost lab-on-a-chip devices. PMID:27479470

  20. DNA Adductomics

    PubMed Central

    2015-01-01

    Systems toxicology is a broad-based approach to describe many of the toxicological features that occur within a living system under stress or subjected to exogenous or endogenous exposures. The ultimate goal is to capture an overview of all exposures and the ensuing biological responses of the body. The term exposome has been employed to refer to the totality of all exposures, and systems toxicology investigates how the exposome influences health effects and consequences of exposures over a lifetime. The tools to advance systems toxicology include high-throughput transcriptomics, proteomics, metabolomics, and adductomics, which is still in its infancy. A well-established methodology for the comprehensive measurement of DNA damage resulting from every day exposures is not fully developed. During the past several decades, the 32P-postlabeling technique has been employed to screen the damage to DNA induced by multiple classes of genotoxicants; however, more robust, specific, and quantitative methods have been sought to identify and quantify DNA adducts. Although triple quadrupole and ion trap mass spectrometry, particularly when using multistage scanning (LC–MSn), have shown promise in the field of DNA adductomics, it is anticipated that high-resolution and accurate-mass LC–MSn instrumentation will play a major role in assessing global DNA damage. Targeted adductomics should also benefit greatly from improved triple quadrupole technology. Once the analytical MS methods are fully mature, DNA adductomics along with other -omics tools will contribute greatly to the field of systems toxicology. PMID:24437709

  1. A multi-year survey of stem-end chip defect in chipping potatoes (Solanum tuberosum L.)

    Technology Transfer Automated Retrieval System (TEKTRAN)

    One of the most serious tuber quality concerns of US chip potato growers is stem-end chip defect, which is defined as a localized post-fry discoloration in and adjacent to the vasculature on the stem end portion of potato chips. The incidence and severity of stem-end chip defect vary with growing lo...

  2. Smart sensor chip based on bioMEMS

    NASA Astrophysics Data System (ADS)

    Madan, Rajesh; Kumar, Sandeep; Bagga, Ellis; Bajpai, Ram P.; Bharadwaj, Lalit M.

    2004-03-01

    The smart sensor chip for simultaneous detection of a large number of disease markers is the most recent interest in the field of nanobiotechnology. Potential applications include miniaturized sensors to detect biological agents and diseases, biocompatible and improved systems for drug delivery. They are the simplest biomicroelectromechanical system (BioMEMS) devices that offer a very promising future to the development of novel physical, chemical and biological sensors. They can simultaneously detect a large number of antigens, antibodies, DNA molecules, trace metals, hormones, proteins, gases, microorganisms, toxins, chemical warfare agents, explosives etc. in gaseous, vacuum and liquid medium. Smart sensor chips would be of greater use in intensive care units (ICUs) where multiple disease markers are to be assessed precisely in very less time. These sensors employ highly specific biochemical reactions between complementary biomolecules in the same way that nature has used in our body to detect, diagnose and treat various types of diseases. They have aroused considerable interest because of their high specificity, ultra-high sensitivity, simplicity, low cost, less analyte requirement (in μl), less steps involved, non-hazardous procedure, quick response, low power requirement and a unique capability of detecting a large number of analytes simultaneously in a single step.

  3. Radiation resistance of sequencing chips for in situ life detection.

    PubMed

    Carr, Christopher E; Rowedder, Holli; Lui, Clarissa S; Zlatkovsky, Ilya; Papalias, Chris W; Bolander, Jarie; Myers, Jason W; Bustillo, James; Rothberg, Jonathan M; Zuber, Maria T; Ruvkun, Gary

    2013-06-01

    Life beyond Earth may be based on RNA or DNA if such life is related to life on Earth through shared ancestry due to meteoritic exchange, such as may be the case for Mars, or if delivery of similar building blocks to habitable environments has biased the evolution of life toward utilizing nucleic acids. In this case, in situ sequencing is a powerful approach to identify and characterize such life without the limitations or expense of returning samples to Earth, and can monitor forward contamination. A new semiconductor sequencing technology based on sensing hydrogen ions released during nucleotide incorporation can enable massively parallel sequencing in a small, robust, optics-free CMOS chip format. We demonstrate that these sequencing chips survive several analogues of space radiation at doses consistent with a 2-year Mars mission, including protons with solar particle event-distributed energy levels and 1 GeV oxygen and iron ions. We find no measurable impact of irradiation at 1 and 5 Gy doses on sequencing quality nor on low-level hardware characteristics. Further testing is required to study the impacts of soft errors as well as to characterize performance under neutron and gamma irradiation and at higher doses, which would be expected during operation in environments with significant trapped energetic particles such as during a mission to Europa. Our results support future efforts to use in situ sequencing to test theories of panspermia and/or whether life has a common chemical basis.

  4. Radiation resistance of sequencing chips for in situ life detection.

    PubMed

    Carr, Christopher E; Rowedder, Holli; Lui, Clarissa S; Zlatkovsky, Ilya; Papalias, Chris W; Bolander, Jarie; Myers, Jason W; Bustillo, James; Rothberg, Jonathan M; Zuber, Maria T; Ruvkun, Gary

    2013-06-01

    Life beyond Earth may be based on RNA or DNA if such life is related to life on Earth through shared ancestry due to meteoritic exchange, such as may be the case for Mars, or if delivery of similar building blocks to habitable environments has biased the evolution of life toward utilizing nucleic acids. In this case, in situ sequencing is a powerful approach to identify and characterize such life without the limitations or expense of returning samples to Earth, and can monitor forward contamination. A new semiconductor sequencing technology based on sensing hydrogen ions released during nucleotide incorporation can enable massively parallel sequencing in a small, robust, optics-free CMOS chip format. We demonstrate that these sequencing chips survive several analogues of space radiation at doses consistent with a 2-year Mars mission, including protons with solar particle event-distributed energy levels and 1 GeV oxygen and iron ions. We find no measurable impact of irradiation at 1 and 5 Gy doses on sequencing quality nor on low-level hardware characteristics. Further testing is required to study the impacts of soft errors as well as to characterize performance under neutron and gamma irradiation and at higher doses, which would be expected during operation in environments with significant trapped energetic particles such as during a mission to Europa. Our results support future efforts to use in situ sequencing to test theories of panspermia and/or whether life has a common chemical basis. PMID:23734755

  5. What Is Mitochondrial DNA?

    MedlinePlus

    ... DNA What is mitochondrial DNA? What is mitochondrial DNA? Although most DNA is packaged in chromosomes within ... proteins. For more information about mitochondria and mitochondrial DNA: Molecular Expressions, a web site from the Florida ...

  6. THE MELTING MECHANISM OF DNA TETHERED TO A SURFACE

    PubMed Central

    QAMHIEH, KHAWLA; WONG, KA-YIU; LYNCH, GILLIAN C.; PETTITT, B. MONTGOMERY

    2009-01-01

    The details of melting of DNA immobilized on a chip or nanoparticle determines the sensitivity and operating characteristics of many analytical and synthetic biotechnological devices. Yet, little is known about the differences in how the DNA melting occurs between a homogeneous solution and that on a chip. We used molecular dynamics simulations to explore possible pathways for DNA melting on a chip. Simulation conditions were chosen to ensure that melting occurred in a submicrosecond timescale. The temperature was set to 400 K and the NaCl concentration was set to 0.1 M. We found less symmetry than in the solution case where for oligomeric double-stranded nucleic acids both ends melted with roughly equal probability. On a prepared silica surface we found melting is dominated by fraying from the end away from the surface. Strand separation was hindered by nonspecific surface adsorption at this temperature. At elevated temperatures the melted DNA was attracted to even uncharged organically coated surfaces demonstrating surface fouling. While hybridization is not the simple reverse of melting, this simulation has implications for the kinetics of hybridization. PMID:19802357

  7. Attachment method for stacked integrated circuit (IC) chips

    DOEpatents

    Bernhardt, A.F.; Malba, V.

    1999-08-03

    An attachment method for stacked integrated circuit (IC) chips is disclosed. The method involves connecting stacked chips, such as DRAM memory chips, to each other and/or to a circuit board. Pads on the individual chips are rerouted to form pads on the side of the chip, after which the chips are stacked on top of each other whereby desired interconnections to other chips or a circuit board can be accomplished via the side-located pads. The pads on the side of a chip are connected to metal lines on a flexible plastic tape (flex) by anisotropically conductive adhesive (ACA). Metal lines on the flex are likewise connected to other pads on chips and/or to pads on a circuit board. In the case of a stack of DRAM chips, pads to corresponding address lines on the various chips may be connected to the same metal line on the flex to form an address bus. This method has the advantage of reducing the number of connections required to be made to the circuit board due to bussing; the flex can accommodate dimensional variation in the alignment of chips in the stack; bonding of the ACA is accomplished at low temperature and is otherwise simpler and less expensive than solder bonding; chips can be bonded to the ACA all at once if the sides of the chips are substantially coplanar, as in the case for stacks of identical chips, such as DRAM. 12 figs.

  8. Attachment method for stacked integrated circuit (IC) chips

    DOEpatents

    Bernhardt, Anthony F.; Malba, Vincent

    1999-01-01

    An attachment method for stacked integrated circuit (IC) chips. The method involves connecting stacked chips, such as DRAM memory chips, to each other and/or to a circuit board. Pads on the individual chips are rerouted to form pads on the side of the chip, after which the chips are stacked on top of each other whereby desired interconnections to other chips or a circuit board can be accomplished via the side-located pads. The pads on the side of a chip are connected to metal lines on a flexible plastic tape (flex) by anisotropically conductive adhesive (ACA). Metal lines on the flex are likewise connected to other pads on chips and/or to pads on a circuit board. In the case of a stack of DRAM chips, pads to corresponding address lines on the various chips may be connected to the same metal line on the flex to form an address bus. This method has the advantage of reducing the number of connections required to be made to the circuit board due to bussing; the flex can accommodate dimensional variation in the alignment of chips in the stack; bonding of the ACA is accomplished at low temperature and is otherwise simpler and less expensive than solder bonding; chips can be bonded to the ACA all at once if the sides of the chips are substantially coplanar, as in the case for stacks of identical chips, such as DRAM.

  9. DNA Microarray-Based Diagnostics.

    PubMed

    Marzancola, Mahsa Gharibi; Sedighi, Abootaleb; Li, Paul C H

    2016-01-01

    The DNA microarray technology is currently a useful biomedical tool which has been developed for a variety of diagnostic applications. However, the development pathway has not been smooth and the technology has faced some challenges. The reliability of the microarray data and also the clinical utility of the results in the early days were criticized. These criticisms added to the severe competition from other techniques, such as next-generation sequencing (NGS), impacting the growth of microarray-based tests in the molecular diagnostic market.Thanks to the advances in the underlying technologies as well as the tremendous effort offered by the research community and commercial vendors, these challenges have mostly been addressed. Nowadays, the microarray platform has achieved sufficient standardization and method validation as well as efficient probe printing, liquid handling and signal visualization. Integration of various steps of the microarray assay into a harmonized and miniaturized handheld lab-on-a-chip (LOC) device has been a goal for the microarray community. In this respect, notable progress has been achieved in coupling the DNA microarray with the liquid manipulation microsystem as well as the supporting subsystem that will generate the stand-alone LOC device.In this chapter, we discuss the major challenges that microarray technology has faced in its almost two decades of development and also describe the solutions to overcome the challenges. In addition, we review the advancements of the technology, especially the progress toward developing the LOC devices for DNA diagnostic applications.

  10. Ultrasensitive FRET-based DNA sensor using PNA/DNA hybridization.

    PubMed

    Yang, Lan-Hee; Ahn, Dong June; Koo, Eunhae

    2016-12-01

    In the diagnosis of genetic diseases, rapid and highly sensitive DNA detection is crucial. Therefore, many strategies for detecting target DNA have been developed, including electrical, optical, and mechanical methods. Herein, a highly sensitive FRET based sensor was developed by using PNA (Peptide Nucleic Acid) probe and QD, in which red color QDs are hybridized with capture probes, reporter probes and target DNAs by EDC-NHS coupling. The hybridized probe with target DNA gives off fluorescent signal due to the energy transfer from QD to Cy5 dye in the reporter probe. Compared to the conventional DNA sensor using DNA probes, the DNA sensor using PNA probes shows higher FRET factor and efficiency due to the higher reactivity between PNA and target DNA. In addition, to elicit the effect of the distance between the donor and the acceptor, we have investigated two types of the reporter probes having Cy5 dyes attached at the different positions of the reporter probes. Results show that the shorter the distance between QDs and Cy5s, the stronger the signal intensity. Furthermore, based on the fluorescence microscopy images using microcapillary chips, the FRET signal is enhanced to be up to 276% times stronger than the signal obtained using the cuvette by the fluorescence spectrometer. These results suggest that the PNA probe system conjugated with QDs can be used as ultrasensitive DNA nanosensors.

  11. Ultrasensitive FRET-based DNA sensor using PNA/DNA hybridization.

    PubMed

    Yang, Lan-Hee; Ahn, Dong June; Koo, Eunhae

    2016-12-01

    In the diagnosis of genetic diseases, rapid and highly sensitive DNA detection is crucial. Therefore, many strategies for detecting target DNA have been developed, including electrical, optical, and mechanical methods. Herein, a highly sensitive FRET based sensor was developed by using PNA (Peptide Nucleic Acid) probe and QD, in which red color QDs are hybridized with capture probes, reporter probes and target DNAs by EDC-NHS coupling. The hybridized probe with target DNA gives off fluorescent signal due to the energy transfer from QD to Cy5 dye in the reporter probe. Compared to the conventional DNA sensor using DNA probes, the DNA sensor using PNA probes shows higher FRET factor and efficiency due to the higher reactivity between PNA and target DNA. In addition, to elicit the effect of the distance between the donor and the acceptor, we have investigated two types of the reporter probes having Cy5 dyes attached at the different positions of the reporter probes. Results show that the shorter the distance between QDs and Cy5s, the stronger the signal intensity. Furthermore, based on the fluorescence microscopy images using microcapillary chips, the FRET signal is enhanced to be up to 276% times stronger than the signal obtained using the cuvette by the fluorescence spectrometer. These results suggest that the PNA probe system conjugated with QDs can be used as ultrasensitive DNA nanosensors. PMID:27612755

  12. Microchannel cooling of face down bonded chips

    DOEpatents

    Bernhardt, Anthony F.

    1993-01-01

    Microchannel cooling is applied to flip-chip bonded integrated circuits, in a manner which maintains the advantages of flip-chip bonds, while overcoming the difficulties encountered in cooling the chips. The technique is suited to either multichip integrated circuit boards in a plane, or to stacks of circuit boards in a three dimensional interconnect structure. Integrated circuit chips are mounted on a circuit board using flip-chip or control collapse bonds. A microchannel structure is essentially permanently coupled with the back of the chip. A coolant delivery manifold delivers coolant to the microchannel structure, and a seal consisting of a compressible elastomer is provided between the coolant delivery manifold and the microchannel structure. The integrated circuit chip and microchannel structure are connected together to form a replaceable integrated circuit module which can be easily decoupled from the coolant delivery manifold and the circuit board. The coolant supply manifolds may be disposed between the circuit boards in a stack and coupled to supplies of coolant through a side of the stack.

  13. Three dimensional, multi-chip module

    DOEpatents

    Bernhardt, A.F.; Petersen, R.W.

    1993-08-31

    A plurality of multi-chip modules are stacked and bonded around the perimeter by sold-bump bonds to adjacent modules on, for instance, three sides of the perimeter. The fourth side can be used for coolant distribution, for more interconnect structures, or other features, depending on particular design considerations of the chip set. The multi-chip modules comprise a circuit board, having a planarized interconnect structure formed on a first major surface, and integrated circuit chips bonded to the planarized interconnect surface. Around the periphery of each circuit board, long, narrow dummy chips'' are bonded to the finished circuit board to form a perimeter wall. The wall is higher than any of the chips on the circuit board, so that the flat back surface of the board above will only touch the perimeter wall. Module-to-module interconnect is laser-patterned on the sides of the boards and over the perimeter wall in the same way and at the same time that chip to board interconnect may be laser-patterned.

  14. Three dimensional, multi-chip module

    DOEpatents

    Bernhardt, Anthony F.; Petersen, Robert W.

    1993-01-01

    A plurality of multi-chip modules are stacked and bonded around the perimeter by sold-bump bonds to adjacent modules on, for instance, three sides of the perimeter. The fourth side can be used for coolant distribution, for more interconnect structures, or other features, depending on particular design considerations of the chip set. The multi-chip modules comprise a circuit board, having a planarized interconnect structure formed on a first major surface, and integrated circuit chips bonded to the planarized interconnect surface. Around the periphery of each circuit board, long, narrow "dummy chips" are bonded to the finished circuit board to form a perimeter wall. The wall is higher than any of the chips on the circuit board, so that the flat back surface of the board above will only touch the perimeter wall. Module-to-module interconnect is laser-patterned o the sides of the boards and over the perimeter wall in the same way and at the same time that chip to board interconnect may be laser-patterned.

  15. Spatial distribution of predicted transcription factor binding sites in Drosophila ChIP peaks.

    PubMed

    Pettie, Kade P; Dresch, Jacqueline M; Drewell, Robert A

    2016-08-01

    In the development of the Drosophila embryo, gene expression is directed by the sequence-specific interactions of a large network of protein transcription factors (TFs) and DNA cis-regulatory binding sites. Once the identity of the typically 8-10bp binding sites for any given TF has been determined by one of several experimental procedures, the sequences can be represented in a position weight matrix (PWM) and used to predict the location of additional TF binding sites elsewhere in the genome. Often, alignments of large (>200bp) genomic fragments that have been experimentally determined to bind the TF of interest in Chromatin Immunoprecipitation (ChIP) studies are trimmed under the assumption that the majority of the binding sites are located near the center of all the aligned fragments. In this study, ChIP/chip datasets are analyzed using the corresponding PWMs for the well-studied TFs; CAUDAL, HUNCHBACK, KNIRPS and KRUPPEL, to determine the distribution of predicted binding sites. All four TFs are critical regulators of gene expression along the anterio-posterior axis in early Drosophila development. For all four TFs, the ChIP peaks contain multiple binding sites that are broadly distributed across the genomic region represented by the peak, regardless of the prediction stringency criteria used. This result suggests that ChIP peak trimming may exclude functional binding sites from subsequent analyses.

  16. Fermilab Physics Department TVC chip

    SciTech Connect

    Hansen, S.; Cotta-Ramusino, A.

    1990-07-01

    The Electronics Group in the Physics Department at Fermilab has designed and has had produced 20 prototypes of a full custom four channel time to voltage converter using the ES2 direct write 2 {mu}m CMOS process. The actual implementation of the design was performed under contract by ASIC designs Inc. of Naperville, Illinois. Each channel has two hit capability and one level of input buffering: that is, up to four voltages representing time internals can be stored from each input for later ADC conversion. The chip produces an edited list of hits and presents the appropriate analog value on its output for each digital value on its hit address lines. The next hit address and analog voltage in the event is presented in response to an external strobe. One current sum proportional to the number of inputs hit for each input buffer is also provided. The chip has been designed to be used on a fastbus TDC card developed here, but it is our belief that it could be adapted to many TDC applications. 2 refs., 8 figs.

  17. Whole-Teflon microfluidic chips

    PubMed Central

    Ren, Kangning; Dai, Wen; Zhou, Jianhua; Su, Jing; Wu, Hongkai

    2011-01-01

    Although microfluidics has shown exciting potential, its broad applications are significantly limited by drawbacks of the materials used to make them. In this work, we present a convenient strategy for fabricating whole-Teflon microfluidic chips with integrated valves that show outstanding inertness to various chemicals and extreme resistance against all solvents. Compared with other microfluidic materials [e.g., poly(dimethylsiloxane) (PDMS)] the whole-Teflon chip has a few more advantages, such as no absorption of small molecules, little adsorption of biomolecules onto channel walls, and no leaching of residue molecules from the material bulk into the solution in the channel. Various biological cells have been cultured in the whole-Teflon channel. Adherent cells can attach to the channel bottom, spread, and proliferate well in the channels (with similar proliferation rate to the cells in PDMS channels with the same dimensions). The moderately good gas permeability of the Teflon materials makes it suitable to culture cells inside the microchannels for a long time. PMID:21536918

  18. Physiologically relevant organs on chips.

    PubMed

    Yum, Kyungsuk; Hong, Soon Gweon; Healy, Kevin E; Lee, Luke P

    2014-01-01

    Recent advances in integrating microengineering and tissue engineering have generated promising microengineered physiological models for experimental medicine and pharmaceutical research. Here we review the recent development of microengineered physiological systems, or also known as "ogans-on-chips", that reconstitute the physiologically critical features of specific human tissues and organs and their interactions. This technology uses microengineering approaches to construct organ-specific microenvironments, reconstituting tissue structures, tissue-tissue interactions and interfaces, and dynamic mechanical and biochemical stimuli found in specific organs, to direct cells to assemble into functional tissues. We first discuss microengineering approaches to reproduce the key elements of physiologically important, dynamic mechanical microenvironments, biochemical microenvironments, and microarchitectures of specific tissues and organs in microfluidic cell culture systems. This is followed by examples of microengineered individual organ models that incorporate the key elements of physiological microenvironments into single microfluidic cell culture systems to reproduce organ-level functions. Finally, microengineered multiple organ systems that simulate multiple organ interactions to better represent human physiology, including human responses to drugs, is covered in this review. This emerging organs-on-chips technology has the potential to become an alternative to 2D and 3D cell culture and animal models for experimental medicine, human disease modeling, drug development, and toxicology.

  19. CHIPPING FRACTURE RESISTANCE OF DENTURE TOOTH MATERIALS

    PubMed Central

    Quinn, G. D.; Giuseppetti, A. A.; Hoffman, K. H.

    2014-01-01

    Objective The applicability of the edge chipping method to denture tooth materials was assessed. These are softer materials than those usually tested by edge chipping. The edge chipping fracture resistances of polymethylmethacrylate (PMMA) based and two filled resin composite denture tooth materials were compared. Methods An edge chipping machine was used to chip rectangular blocks and flattened anterior denture teeth. Force versus edge distance data were collected over a broad range of forces and distances. Between 20 and 65 chips were made per condition depending upon the material, the scatter, and the indenter type. Different indenter types were used including Rockwell C, sharp conical 120°, Knoop, and Vickers. The edge toughness, Te, was evaluated for different indenter types. Results The edge chipping data collected on the blocks matched the data collected from flattened teeth. High scatter, particularly at large distances and loads, meant that many tests (up to 64) were necessary to compare the denture tooth materials and to ascertain the appropriate data trends. A linear force – distance trend analysis was adequate for comparing these materials. A power law trend might be more appropriate, but the large scatter obscured the definitive determination of the precise trend. Different indenters produce different linear trends, with the ranking of: sharp conical 120°, Rockwell C, and Knoop, from lowest to highest edge toughness. Vickers indenter data were extremely scattered and a sensible trend could not be obtained. Edge toughness was inversely correlated to hardness. Significance Edge chipping data collected either from simple laboratory scale test blocks or from actual denture teeth may be used to evaluate denture materials. The edge chipping method’s applicability has been extended to another class of restorative materials. PMID:24674342

  20. Suspended Solid-state Membranes on Glass Chips with Sub 1-pF Capacitance for Biomolecule Sensing Applications

    PubMed Central

    Balan, Adrian; Chien, Chen-Chi; Engelke, Rebecca; Drndić, Marija

    2015-01-01

    Solid-state membranes are finding use in many applications in nanoelectronics and nanomedicine, from single molecule sensors to water filtration, and yet many of their electronics applications are limited by the relatively high current noise and low bandwidth stemming from the relatively high capacitance (>10 pF) of the membrane chips. To address this problem, we devised an integrated fabrication process to grow and define circular silicon nitride membranes on glass chips that successfully lower the chip capacitance to below 1 pF. We use these devices to demonstrate low-noise, high-bandwidth DNA translocation measurements. We also make use of this versatile, low-capacitance platform to suspend other thin, two-dimensional membrane such as graphene. PMID:26644307

  1. Suspended Solid-state Membranes on Glass Chips with Sub 1-pF Capacitance for Biomolecule Sensing Applications

    NASA Astrophysics Data System (ADS)

    Balan, Adrian; Chien, Chen-Chi; Engelke, Rebecca; Drndić, Marija

    2015-12-01

    Solid-state membranes are finding use in many applications in nanoelectronics and nanomedicine, from single molecule sensors to water filtration, and yet many of their electronics applications are limited by the relatively high current noise and low bandwidth stemming from the relatively high capacitance (>10 pF) of the membrane chips. To address this problem, we devised an integrated fabrication process to grow and define circular silicon nitride membranes on glass chips that successfully lower the chip capacitance to below 1 pF. We use these devices to demonstrate low-noise, high-bandwidth DNA translocation measurements. We also make use of this versatile, low-capacitance platform to suspend other thin, two-dimensional membrane such as graphene.

  2. Suspended Solid-state Membranes on Glass Chips with Sub 1-pF Capacitance for Biomolecule Sensing Applications

    NASA Astrophysics Data System (ADS)

    Chien, Chen-Chi; Balan, Adrian; Engelke, Rebecca; Drndic, Marija

    Solid-state membranes are finding use in many applications in nanoelectronics and nanomedicine, from single molecule sensors to water filtration, and yet many of their electronics applications are limited by the current noise and low bandwidth stemming from the relatively high capacitance (more than 10 pF) of the membrane chips. To address this problem, we devised an integrated fabrication process to grow and define circular silicon nitride membranes on glass chips that successfully lower the chip capacitance to below 1 pF. We use these devices to demonstrate low-noise, high-bandwidth DNA translocation measurements. We also make use of this versatile, low-capacitance platform to suspend other thin, two-dimensional membranes such as graphene.

  3. Suspended Solid-state Membranes on Glass Chips with Sub 1-pF Capacitance for Biomolecule Sensing Applications.

    PubMed

    Balan, Adrian; Chien, Chen-Chi; Engelke, Rebecca; Drndić, Marija

    2015-12-08

    Solid-state membranes are finding use in many applications in nanoelectronics and nanomedicine, from single molecule sensors to water filtration, and yet many of their electronics applications are limited by the relatively high current noise and low bandwidth stemming from the relatively high capacitance (>10 pF) of the membrane chips. To address this problem, we devised an integrated fabrication process to grow and define circular silicon nitride membranes on glass chips that successfully lower the chip capacitance to below 1 pF. We use these devices to demonstrate low-noise, high-bandwidth DNA translocation measurements. We also make use of this versatile, low-capacitance platform to suspend other thin, two-dimensional membrane such as graphene.

  4. Ancient DNA.

    PubMed

    Willerslev, Eske; Cooper, Alan

    2005-01-01

    In the past two decades, ancient DNA research has progressed from the retrieval of small fragments of mitochondrial DNA from a few late Holocene specimens, to large-scale studies of ancient populations, phenotypically important nuclear loci, and even whole mitochondrial genome sequences of extinct species. However, the field is still regularly marred by erroneous reports, which underestimate the extent of contamination within laboratories and samples themselves. An improved understanding of these processes and the effects of damage on ancient DNA templates has started to provide a more robust basis for research. Recent methodological advances have included the characterization of Pleistocene mammal populations and discoveries of DNA preserved in ancient sediments. Increasingly, ancient genetic information is providing a unique means to test assumptions used in evolutionary and population genetics studies to reconstruct the past. Initial results have revealed surprisingly complex population histories, and indicate that modern phylogeographic studies may give misleading impressions about even the recent evolutionary past. With the advent and uptake of appropriate methodologies, ancient DNA is now positioned to become a powerful tool in biological research and is also evolving new and unexpected uses, such as in the search for extinct or extant life in the deep biosphere and on other planets.

  5. Ancient DNA

    PubMed Central

    Willerslev, Eske; Cooper, Alan

    2004-01-01

    In the past two decades, ancient DNA research has progressed from the retrieval of small fragments of mitochondrial DNA from a few late Holocene specimens, to large-scale studies of ancient populations, phenotypically important nuclear loci, and even whole mitochondrial genome sequences of extinct species. However, the field is still regularly marred by erroneous reports, which underestimate the extent of contamination within laboratories and samples themselves. An improved understanding of these processes and the effects of damage on ancient DNA templates has started to provide a more robust basis for research. Recent methodological advances have included the characterization of Pleistocene mammal populations and discoveries of DNA preserved in ancient sediments. Increasingly, ancient genetic information is providing a unique means to test assumptions used in evolutionary and population genetics studies to reconstruct the past. Initial results have revealed surprisingly complex population histories, and indicate that modern phylogeographic studies may give misleading impressions about even the recent evolutionary past. With the advent and uptake of appropriate methodologies, ancient DNA is now positioned to become a powerful tool in biological research and is also evolving new and unexpected uses, such as in the search for extinct or extant life in the deep biosphere and on other planets. PMID:15875564

  6. DNA vaccines

    NASA Astrophysics Data System (ADS)

    Gregersen, Jens-Peter

    2001-12-01

    Immunization by genes encoding immunogens, rather than with the immunogen itself, has opened up new possibilities for vaccine research and development and offers chances for new applications and indications for future vaccines. The underlying mechanisms of antigen processing, immune presentation and regulation of immune responses raise high expectations for new and more effective prophylactic or therapeutic vaccines, particularly for vaccines against chronic or persistent infectious diseases and tumors. Our current knowledge and experience of DNA vaccination is summarized and critically reviewed with particular attention to basic immunological mechanisms, the construction of plasmids, screening for protective immunogens to be encoded by these plasmids, modes of application, pharmacokinetics, safety and immunotoxicological aspects. DNA vaccines have the potential to accelerate the research phase of new vaccines and to improve the chances of success, since finding new immunogens with the desired properties is at least technically less demanding than for conventional vaccines. However, on the way to innovative vaccine products, several hurdles have to be overcome. The efficacy of DNA vaccines in humans appears to be much less than indicated by early studies in mice. Open questions remain concerning the persistence and distribution of inoculated plasmid DNA in vivo, its potential to express antigens inappropriately, or the potentially deleterious ability to insert genes into the host cell's genome. Furthermore, the possibility of inducing immunotolerance or autoimmune diseases also needs to be investigated more thoroughly, in order to arrive at a well-founded consensus, which justifies the widespread application of DNA vaccines in a healthy population.

  7. DNA topoisomerases.

    PubMed

    Wang, J C

    1996-01-01

    The various problems of disentangling DNA strands or duplexes in a cell are all rooted in the double-helical structure of DNA. Three distinct subfamilies of enzymes, known as the DNA topoisomerases, have evolved to solve these problems. This review focuses on work in the past decade on the mechanisms and cellular functions of these enzymes. Newly discovered members and recent biochemical and structural results are reviewed, and mechanistic implications of these results are summarized. The primary cellular functions of these enzymes, including their roles in replication, transcription, chromosome condensation, and the maintenance of genome stability, are then discussed. The review ends with a summary of the regulation of the cellular levels of these enzymes and a discussion of their association with other cellular proteins.

  8. Voltage Regulator Chip: Power Supplies on a Chip

    SciTech Connect

    2010-09-01

    ADEPT Project: CPES at Virginia Tech is finding ways to save real estate on a computer's motherboard that could be used for other critical functions. Every computer processor today contains a voltage regulator that automatically maintains a constant level of electricity entering the device. These regulators contain bulky components and take up about 30% of a computer's motherboard. CPES at Virginia Tech is developing a voltage regulator that uses semiconductors made of gallium nitride on silicon (GaN-on-Si) and high-frequency soft magnetic material. These materials are integrated on a small, 3D chip that can handle the same amount of power as traditional voltage regulators at 1/10 the size and with improved efficiency. The small size also frees up to 90% of the motherboard space occupied by current voltage regulators.

  9. Improvement of Temperature Uniformity for Polymerase Chain Reaction Chip with Heat Spreader

    NASA Astrophysics Data System (ADS)

    Chen, Rong-Sheng; Mao, Chao-Yang; Chen, Yung-Shieng

    2007-11-01

    For polymerase chain reaction (PCR) applications, a uniform temperature field in the microreactor is crucial. In this paper, we report on the electrothermal and computational fluid dynamics (CFD) simulations performed with the aim of optimizing the temperature distribution by heat spreaders for PCR application. Firstly, the equivalent resistivity of the microresistor heater is evaluated, and a conformable result is then verified by comparing with the experimental result using a prototype PCR chip. Secondly, the temperature distribution at 94 °C in the PCR chip is investigated. Furthermore, a heat spreader is inserted into the PCR chip to reduce the temperature difference in the DNA sample and thus improve the temperature uniformity effectively. The results demonstrated that the effective volume percentage and the energy consumption in the chamber are positively related to the thickness of the heat spreader, while the temperature difference is inversely related to the thickness of the heat spreader. Finally, the (b)-design is better than the (a)-design in terms of both the increase in effective volume percentage of the DNA sample and the decrease in energy consumption. In other words, the (b)-design is recognized as having better temperature uniformity.

  10. Health hazards caused by fungi in stored wood chips

    SciTech Connect

    Thoernquist, T.; Lundstroem, H.

    1982-11-01

    In connection with using wood chips for fuel in heating buildings, a number of people in Sweden were taken ill with a respiratory allergy similar to wood trimmer's disease and farmer's lung. The disease is presumably caused by airborne fungal particles (spores and hyphae) which are inhaled when working with infected wood chips. The occurrence of fungal particles in the air in wood chip storage rooms, halls, and kitchens was studied in 64 buildings heated by chips. Sampling was carried out by exposing 9-cm petri dishes containing malt agar. In the chip storage rooms of 10 of the 64 buildings examined, more than 500 fungal colonies were recorded before disturbing the chips. After disturbance the number of buildings with more than 500 colonies increased to 28. In the halls in three of the buildings and in the kitchens of two, more than 500 fungal colonies were recorded. The number of fungal particles in wood chip storage is mainly dependent on the condition of the raw material before chipping, tree species, and the final storage period. To reduce the risk of large numbers of fungal particles in stored chips, the trees should be limbed before chipping and the stems preferably dried. Hardwood chips are more easily infected by fungi than chips of coniferous wood. The storage of wood chips for periods longer than 3 months should be avoided and a Class 2B protective mask should always be worn when handling chips feared to be infected by fungi. (Refs. 5).

  11. Digital isothermal quantification of nucleic acids via simultaneous chemical initiation of recombinase polymerase amplification reactions on SlipChip.

    PubMed

    Shen, Feng; Davydova, Elena K; Du, Wenbin; Kreutz, Jason E; Piepenburg, Olaf; Ismagilov, Rustem F

    2011-05-01

    In this paper, digital quantitative detection of nucleic acids was achieved at the single-molecule level by chemical initiation of over one thousand sequence-specific, nanoliter isothermal amplification reactions in parallel. Digital polymerase chain reaction (digital PCR), a method used for quantification of nucleic acids, counts the presence or absence of amplification of individual molecules. However, it still requires temperature cycling, which is undesirable under resource-limited conditions. This makes isothermal methods for nucleic acid amplification, such as recombinase polymerase amplification (RPA), more attractive. A microfluidic digital RPA SlipChip is described here for simultaneous initiation of over one thousand nL-scale RPA reactions by adding a chemical initiator to each reaction compartment with a simple slipping step after instrument-free pipet loading. Two designs of the SlipChip, two-step slipping and one-step slipping, were validated using digital RPA. By using the digital RPA SlipChip, false-positive results from preinitiation of the RPA amplification reaction before incubation were eliminated. End point fluorescence readout was used for "yes or no" digital quantification. The performance of digital RPA in a SlipChip was validated by amplifying and counting single molecules of the target nucleic acid, methicillin-resistant Staphylococcus aureus (MRSA) genomic DNA. The digital RPA on SlipChip was also tolerant to fluctuations of the incubation temperature (37-42 °C), and its performance was comparable to digital PCR on the same SlipChip design. The digital RPA SlipChip provides a simple method to quantify nucleic acids without requiring thermal cycling or kinetic measurements, with potential applications in diagnostics and environmental monitoring under resource-limited settings. The ability to initiate thousands of chemical reactions in parallel on the nanoliter scale using solvent-resistant glass devices is likely to be useful for a broader

  12. Disposable polyester-toner electrophoresis microchips for DNA analysis.

    PubMed

    Duarte, Gabriela R M; Coltro, Wendell K T; Borba, Juliane C; Price, Carol W; Landers, James P; Carrilho, Emanuel

    2012-06-01

    Microchip electrophoresis has become a powerful tool for DNA separation, offering all of the advantages typically associated with miniaturized techniques: high speed, high resolution, ease of automation, and great versatility for both routine and research applications. Various substrate materials have been used to produce microchips for DNA separations, including conventional (glass, silicon, and quartz) and alternative (polymers) platforms. In this study, we perform DNA separation in a simple and low-cost polyester-toner (PeT)-based electrophoresis microchip. PeT devices were fabricated by a direct-printing process using a 600 dpi-resolution laser printer. DNA separations were performed on PeT chip with channels filled with polymer solutions (0.5% m/v hydroxyethylcellulose or hydroxypropylcellulose) at electric fields ranging from 100 to 300 V cm(-1). Separation of DNA fragments between 100 and 1000 bp, with good correlation of the size of DNA fragments and mobility, was achieved in this system. Although the mobility increased with increasing electric field, separations showed the same profile regardless of the electric field. The system provided good separation efficiency (215,000 plates per m for the 500 bp fragment) and the separation was completed in 4 min for 1000 bp fragment ladder. The cost of a given chip is approximately $0.15 and it takes less than 10 minutes to prepare a single device.

  13. IC chip stress during plastic package molding

    SciTech Connect

    Palmer, D.W.; Benson, D.A.; Peterson, D.W.; Sweet, J.N.

    1998-02-01

    Approximately 95% of the world`s integrated chips are packaged using a hot, high pressure transfer molding process. The stress created by the flow of silica powder loaded epoxy can displace the fine bonding wires and can even distort the metalization patterns under the protective chip passivation layer. In this study the authors developed a technique to measure the mechanical stress over the surface of an integrated circuit during the molding process. A CMOS test chip with 25 diffused resistor stress sensors was applied to a commercial lead frame. Both compression and shear stresses were measured at all 25 locations on the surface of the chip every 50 milliseconds during molding. These measurements have a fine time and stress resolution which should allow comparison with computer simulation of the molding process, thus allowing optimization of both the manufacturing process and mold geometry.

  14. Accelerator on a Chip: How It Works

    SciTech Connect

    2014-06-30

    In an advance that could dramatically shrink particle accelerators for science and medicine, researchers used a laser to accelerate electrons at a rate 10 times higher than conventional technology in a nanostructured glass chip smaller than a grain of rice.

  15. DNA Methylation

    PubMed Central

    Marinus, M.G.; Løbner-Olesen, A.

    2014-01-01

    The DNA of E. coli contains 19,120 6-methyladenines and 12,045 5-methylcytosines in addition to the four regular bases and these are formed by the postreplicative action of three DNA methyltransferases. The majority of the methylated bases are formed by the Dam and Dcm methyltransferases encoded by the dam (DNA adenine methyltransferase) and dcm (DNA cytosine methyltransferase) genes. Although not essential, Dam methylation is important for strand discrimination during repair of replication errors, controlling the frequency of initiation of chromosome replication at oriC, and regulation of transcription initiation at promoters containing GATC sequences. In contrast, there is no known function for Dcm methylation although Dcm recognition sites constitute sequence motifs for Very Short Patch repair of T/G base mismatches. In certain bacteria (e.g., Vibrio cholerae, Caulobacter crescentus) adenine methylation is essential and in C. crescentus, it is important for temporal gene expression which, in turn, is required for coordinating chromosome initiation, replication and division. In practical terms, Dam and Dcm methylation can inhibit restriction enzyme cleavage; decrease transformation frequency in certain bacteria; decrease the stability of short direct repeats; are necessary for site-directed mutagenesis; and to probe eukaryotic structure and function. PMID:26442938

  16. DNA Investigations.

    ERIC Educational Resources Information Center

    Mayo, Ellen S.; Bertino, Anthony J.

    1991-01-01

    Presents a simulation activity that allow students to work through the exercise of DNA profiling and to grapple with some analytical and ethical questions involving a couple arranging with a surrogate mother to have a baby. Can be used to teach the principles of restriction enzyme digestion, gel electrophoresis, and probe hybridization. (MDH)

  17. Design and Validation of DNA Libraries for Multiplexing Proximity Ligation Assays

    PubMed Central

    Gobet, Nicolas; Ketterer, Simon; Meier, Matthias

    2014-01-01

    Here, we present an in silico, analytical procedure for designing and testing orthogonal DNA templates for multiplexing of the proximity ligation assay (PLA). PLA is a technology for the detection of protein interactions, post-translational modifications, and protein concentrations. To enable multiplexing of the PLA, the target information of antibodies was encoded within the DNA template of a PLA, where each template comprised four single-stranded DNA molecules. Our DNA design procedure followed the principles of minimizing the free energy of DNA cross-hybridization. To validate the functionality, orthogonality, and efficiency of the constructed template libraries, we developed a high-throughput solid-phase rolling-circle amplification assay and solid-phase PLA on a microfluidic platform. Upon integration on a microfluidic chip, 640 miniaturized pull-down assays for oligonucleotides or antibodies could be performed in parallel together with steps of DNA ligation, isothermal amplification, and detection under controlled microenvironments. From a large computed PLA template library, we randomly selected 10 template sets and tested all DNA combinations for cross-reactivity in the presence and absence of antibodies. By using the microfluidic chip application, we determined rapidly the false-positive rate of the design procedure, which was less than 1%. The combined theoretical and experimental procedure is applicable for high-throughput PLA studies on a microfluidic chip. PMID:25386748

  18. [Detection of transgenic crop with gene chip].

    PubMed

    Huang, Ying-Chun; Sun, Chun-Yun; Feng, Hong; Hu, Xiao-Dong; Yin, Hai-Bin

    2003-05-01

    Some selected available sequences of reporter genes,resistant genes, promoters and terminators are amplified by PCR for the probes of transgenic crop detection gene chip. These probes are arrayed at definite density and printed on the surface of amino-slides by bioRobot MicroGrid II. Results showed that gene chip worked quickly and correctly, when transgenic rice, pawpaw,maize and soybean were applied. PMID:15639876

  19. Isolation of the cDNA for erythrocyte integral membrane protein of 28 kilodaltons: member of an ancient channel family.

    PubMed Central

    Preston, G M; Agre, P

    1991-01-01

    CHIP28 is a 28-kDa integral membrane protein with similarities to membrane channels and is found in erythrocytes and renal tubules. A cDNA for CHIP28 was isolated from human fetal liver cDNA template by a three-step polymerase chain reaction (PCR) cloning strategy, starting with degenerate oligonucleotide primers corresponding to the N-terminal amino acid sequence determined from purified CHIP28 protein. Using the third-step PCR product as a probe, we isolated a recombinant from a human bone marrow cDNA library. The combined sequence of the PCR products and bone marrow cDNA contains 38 base pairs of 5' untranslated nucleotide sequence, an 807-bp open reading frame, and approximately 2 kilobases of 3' untranslated sequence containing a polyadenylation signal. This corresponds to the 3.1-kilobase transcript identified by RNA blot-hybridization analysis. Authenticity of the deduced amino acid sequence of the CHIP28 protein C terminus was confirmed by expression and immunoblotting. Analysis of the deduced amino acid sequence suggests that CHIP28 protein contains six bilayer-spanning domains, two exofacial potential N-glycosylation sites, and intracellular N and C termini. Search of the DNA sequence data base revealed a strong homology with the major intrinsic protein of bovine lens, which is the prototype of an ancient but recently recognized family of membrane channels. These proteins are believed to form channels permeable to water and possibly other small molecules. CHIP28 shares homology with all known members of this channel family, and it is speculated that CHIP28 has a similar function. Images PMID:1722319

  20. RisaAligner software for aligning fluorescence data between Agilent 2100 Bioanalyzer chips: Application to soil microbial community analysis.

    PubMed

    Navarro, Elisabeth; Fabrègue, Olivier; Scorretti, Riccardo; Reboulet, Jérémy; Simonet, Pascal; Dawson, Lorna; Demanèche, Sandrine

    2015-12-01

    Ribosomal Intergenic Spacer Analysis (RISA) is a high-resolution and highly reproducible fingerprinting technique for discriminating between microbial communities. The community profiles can be visualized using the Agilent 2100 Bioanalyzer. Comparison between fingerprints relies upon precise estimation of all amplified DNA fragment lengths; however, size standard computation can vary between gel runs. For complex samples such as soil microbial communities, discrimination by fragment size is not always sufficient. In such cases, the comparison of whole fluorescence data as a function of time (electrophoregrams) is more appropriate. When electrophoregrams [fluorescence = f (time)] are used, and more than one chip is involved, electrophoregram comparisons are challenging due to experimental variations between chips and the lack of correction by the Agilent software in such situations. Here we present RisaAligner software for analyzing and comparing electrophoregrams from Agilent chips using a nonlinear ladder-alignment algorithm. We demonstrate the robustness and substantial improvement of data analysis by analyzing soil microbial profiles obtained with Agilent DNA 1000 and High Sensitivity chips.

  1. RisaAligner software for aligning fluorescence data between Agilent 2100 Bioanalyzer chips: Application to soil microbial community analysis.

    PubMed

    Navarro, Elisabeth; Fabrègue, Olivier; Scorretti, Riccardo; Reboulet, Jérémy; Simonet, Pascal; Dawson, Lorna; Demanèche, Sandrine

    2015-12-01

    Ribosomal Intergenic Spacer Analysis (RISA) is a high-resolution and highly reproducible fingerprinting technique for discriminating between microbial communities. The community profiles can be visualized using the Agilent 2100 Bioanalyzer. Comparison between fingerprints relies upon precise estimation of all amplified DNA fragment lengths; however, size standard computation can vary between gel runs. For complex samples such as soil microbial communities, discrimination by fragment size is not always sufficient. In such cases, the comparison of whole fluorescence data as a function of time (electrophoregrams) is more appropriate. When electrophoregrams [fluorescence = f (time)] are used, and more than one chip is involved, electrophoregram comparisons are challenging due to experimental variations between chips and the lack of correction by the Agilent software in such situations. Here we present RisaAligner software for analyzing and comparing electrophoregrams from Agilent chips using a nonlinear ladder-alignment algorithm. We demonstrate the robustness and substantial improvement of data analysis by analyzing soil microbial profiles obtained with Agilent DNA 1000 and High Sensitivity chips. PMID:26651514

  2. Viscosimeter on a microfluidic chip.

    PubMed

    Guillot, Pierre; Panizza, Pascal; Salmon, Jean-Baptiste; Joanicot, Mathieu; Colin, Annie; Bruneau, Charles-Henri; Colin, Thierry

    2006-07-01

    In this work, a viscosimeter implemented on a microfluidic chip is presented. The physical principle of this system is to use laminar parallel flows in a microfluidic channel. The fluid to be studied flows side by side with a reference fluid of known viscosity. By using optical microscopy, the shape of the interface between both fluids can be determined. Knowing the flow rates of the two liquids and the geometrical features of the channel, the mean shear rate sustained by the fluid and its viscosity can thus be computed. Accurate and precise measurements of the viscosity as a function of the shear rate can be made using less than 300 microL of fluid. Several complex fluids are tested with viscosities ranging from 10(-)(3) to 70 Pa.s.

  3. BLOOD-ON-A-CHIP

    PubMed Central

    Toner, Mehmet; Irimia, Daniel

    2013-01-01

    Accurate, fast, and affordable analysis of the cellular component of blood is of prime interest for medicine and research. Yet, most often sample preparation procedures for blood analysis involve handling steps prone to introducing artifacts, whereas analysis methods commonly require skilled technicians and well-equipped, expensive laboratories. Developing more gentle protocols and affordable instruments for specific blood analysis tasks is becoming possible through the recent progress in the area of microfluidics and lab-on-a-chip-type devices. Precise control over the cell microenvironment during separation procedures and the ability to scale down the analysis to very small volumes of blood are among the most attractive capabilities of the new approaches. Here we review some of the emerging principles for manipulating blood cells at microscale and promising high-throughput approaches to blood cell separation using microdevices. Examples of specific single-purpose devices are described together with integration strategies for blood cell separation and analysis modules. PMID:16004567

  4. Chip Scale Package Implementation Challenges

    NASA Technical Reports Server (NTRS)

    Ghaffarian, Reza

    1998-01-01

    The JPL-led MicrotypeBGA Consortium of enterprises representing government agencies and private companies have jointed together to pool in-kind resources for developing the quality and reliability of chip scale packages (CSPs) for a variety of projects. In the process of building the Consortium CSP test vehicles, many challenges were identified regarding various aspects of technology implementation. This paper will present our experience in the areas of technology implementation challenges, including design and building both standard and microvia boards, and assembly of two types of test vehicles. We also discuss the most current package isothermal aging to 2,000 hours at 100 C and 125 C and thermal cycling test results to 1,700 cycles in the range of -30 to 100 C.

  5. Advanced atom chips with two metal layers.

    SciTech Connect

    Stevens, James E.; Blain, Matthew Glenn; Benito, Francisco M.; Biedermann, Grant

    2010-12-01

    A design concept, device layout, and monolithic microfabrication processing sequence have been developed for a dual-metal layer atom chip for next-generation positional control of ultracold ensembles of trapped atoms. Atom chips are intriguing systems for precision metrology and quantum information that use ultracold atoms on microfabricated chips. Using magnetic fields generated by current carrying wires, atoms are confined via the Zeeman effect and controllably positioned near optical resonators. Current state-of-the-art atom chips are single-layer or hybrid-integrated multilayer devices with limited flexibility and repeatability. An attractive feature of multi-level metallization is the ability to construct more complicated conductor patterns and thereby realize the complex magnetic potentials necessary for the more precise spatial and temporal control of atoms that is required. Here, we have designed a true, monolithically integrated, planarized, multi-metal-layer atom chip for demonstrating crossed-wire conductor patterns that trap and controllably transport atoms across the chip surface to targets of interest.

  6. On-chip positionable photonic waveguides for chip-to-chip optical interconnects

    NASA Astrophysics Data System (ADS)

    Peters, Tjitte-Jelte; Tichem, Marcel

    2016-05-01

    This paper reports on the progress related to a multichannel photonic alignment concept, aiming for sub-micrometer precision in the alignment of the waveguides of two photonic integrated circuits (PICs). The concept consists of two steps: chip-to-chip positioning and chip bonding provide a coarse alignment after which waveguide-to-waveguide positioning and fixing result in a fine alignment. For the waveguide-to-waveguide alignment, an alignment functionality is developed and integrated in one of the PICs, consisting of mechanically flexible waveguides and MEMS actuators. This paper reports on the fabrication and characterization of a mechanically flexible waveguide array that can be positioned by two out-of-plane actuators. Thermal actuators are integrated with mechanically flexible waveguide beams to enable positioning them with high precision. By adding a poly-Si pattern on top of SiO2 beams, an out-of-plane bimorph actuator can be realized. An analytical model enables estimating the curvature and the deflection of a single bimorph beam. Acquiring a small initial deflection while having a large motion range of the actuator proves to have conflicting demands on the poly-Si/SiO2 thickness ratio. In this paper, we show that suspended waveguide arrays with integrated alignment functionality have an initial deflection- they curl up- due to residual stress in the materials. The actuators can be operated using a driving voltage between 0V to 45V, corresponding to ~50mW. Using higher voltages brings the risk of permanently changing the material properties of the heaters. The actuators can accomplish an out-of-plane crossbar translation up to 6.5 μm at ~50mW as well as a rotation around the propagation direction of the light ranging from -0:1° to 0.1°. At a constant actuation power of ~50mW, the crossbar shows a drift in vertical deflection of 0.16 μm over a time of 30 min.

  7. Microfluidic device for DNA amplification of single cancer cells isolated from whole blood by self-seeding microwells.

    PubMed

    Yang, Yoonsun; Rho, Hoon Suk; Stevens, Michiel; Tibbe, Arjan G J; Gardeniers, Han; Terstappen, Leon W M M

    2015-11-21

    Self-seeding microwell chips can sort single cells into 6400 wells based on cell size and their identity verified by immunofluorescence staining. Here, we developed a microfluidic device in which these single cells can be placed, lysed and their DNA amplified for further interrogation. Whole blood spiked with MCF7 tumor cells was passed through the microwell chips after leukocyte depletion and 37% of the MCF7 cells were identified by epithelial cell adhesion molecule (EpCAM) staining in the microwells. Identified single cells were punched into the reaction chamber of the microfluidic device and reagents for cell lysis and DNA amplification introduced sequentially by peristaltic pumping of micro-valves. On-chip lysis and amplification was performed in 8 parallel chambers yielding a 10,000 fold amplification of DNA. Accessibility of the sample through the reaction chamber allowed for easy retrieval and interrogation of target-specific genes to characterize the tumor cells.

  8. Sample preparation module for bacterial lysis and isolation of DNA from human urine

    PubMed Central

    Gillers, Sara; Zhang, Jane Y.; Singh, Satish; Klapperich, Catherine M.

    2015-01-01

    Silica impregnated polymer monolithic columns may provide a simple method for lysing and extracting DNA from bacteria inside of microfluidic chips. Here we use Escherichia coli as a test organism for a point of care thermoplastic microfluidic module designed to take in a urine sample, mix it with lysis buffer, and perform a hybrid chemical/mechanical lysis and solid phase extraction of nucleic acids from the sample. To demonstrate proof-of-concept, we doped human hematuric urine samples with E. coli at concentrations ranging from 101–105 colony-forming units/mL (CFU/mL) to simulate patient samples. We then performed on-chip lysis and DNA extraction. The bacterial DNA was amplified using real-time PCR demonstrating lysis and isolation down to 101 CFU/mL. Results were comparable to a commercial kit at higher concen trations and performed better at recovering DNA at lower concentrations. PMID:19130239

  9. Knowledge-based image processing for on-off type DNA microarray

    NASA Astrophysics Data System (ADS)

    Kim, Jong D.; Kim, Seo K.; Cho, Jeong S.; Kim, Jongwon

    2002-06-01

    This paper addresses the image processing technique for discriminating whether the probes are hybrized with target DNA in the Human Papilloma Virus (HPV) DNA Chip designed for genotyping HPV. In addition to the probes, the HPV DNA chip has markers that always react with the sample DNA. The positions of probe-dots in the final scanned image are fixed relative to the marker-dot locations with a small variation according to the accuracy of the dotter and the scanner. The probes are duplicated 4 times for the diagnostic stability. The prior knowledges such as the maker relative distance and the duplication information of probes is integrated into the template matching technique with the normalized correlation measure. Results show that the employment of both of the prior knowledges is to simply average the template matching measures over the positions of the markers and probes. The eventual proposed scheme yields stable marker locating and probe classification.

  10. A Nanoscale, Liquid-Phase DNA Separation Device Based on Brownian Ratchets

    NASA Astrophysics Data System (ADS)

    Bader, Joel S.

    1998-03-01

    Realizing the goals of the Human Genome Project depends on the ability to perform size-based separations of DNA molecules. DNA analysis has traditionally required inconvenient gel-based electrophoretic separations. We describe a novel, micromachined, non-electrophoretic device suitable for lab-on-a-chip applications. The device is designed to transport DNA using an asymmetric, periodic potential to rectify Brownian motion. The separation occurs in a homogeneous liquid, avoiding the use of gels or other special media. Experimental results from a working prototype NanoNiagara device validate theoretical predictions of its ability to transport DNA molecules based on size.

  11. Picoliter Well Array Chip-Based Digital Recombinase Polymerase Amplification for Absolute Quantification of Nucleic Acids.

    PubMed

    Li, Zhao; Liu, Yong; Wei, Qingquan; Liu, Yuanjie; Liu, Wenwen; Zhang, Xuelian; Yu, Yude

    2016-01-01

    Absolute, precise quantification methods expand the scope of nucleic acids research and have many practical applications. Digital polymerase chain reaction (dPCR) is a powerful method for nucleic acid detection and absolute quantification. However, it requires thermal cycling and accurate temperature control, which are difficult in resource-limited conditions. Accordingly, isothermal methods, such as recombinase polymerase amplification (RPA), are more attractive. We developed a picoliter well array (PWA) chip with 27,000 consistently sized picoliter reactions (314 pL) for isothermal DNA quantification using digital RPA (dRPA) at 39°C. Sample loading using a scraping liquid blade was simple, fast, and required small reagent volumes (i.e., <20 μL). Passivating the chip surface using a methoxy-PEG-silane agent effectively eliminated cross-contamination during dRPA. Our creative optical design enabled wide-field fluorescence imaging in situ and both end-point and real-time analyses of picoliter wells in a 6-cm(2) area. It was not necessary to use scan shooting and stitch serial small images together. Using this method, we quantified serial dilutions of a Listeria monocytogenes gDNA stock solution from 9 × 10(-1) to 4 × 10(-3) copies per well with an average error of less than 11% (N = 15). Overall dRPA-on-chip processing required less than 30 min, which was a 4-fold decrease compared to dPCR, requiring approximately 2 h. dRPA on the PWA chip provides a simple and highly sensitive method to quantify nucleic acids without thermal cycling or precise micropump/microvalve control. It has applications in fast field analysis and critical clinical diagnostics under resource-limited settings. PMID:27074005

  12. Picoliter Well Array Chip-Based Digital Recombinase Polymerase Amplification for Absolute Quantification of Nucleic Acids.

    PubMed

    Li, Zhao; Liu, Yong; Wei, Qingquan; Liu, Yuanjie; Liu, Wenwen; Zhang, Xuelian; Yu, Yude

    2016-01-01

    Absolute, precise quantification methods expand the scope of nucleic acids research and have many practical applications. Digital polymerase chain reaction (dPCR) is a powerful method for nucleic acid detection and absolute quantification. However, it requires thermal cycling and accurate temperature control, which are difficult in resource-limited conditions. Accordingly, isothermal methods, such as recombinase polymerase amplification (RPA), are more attractive. We developed a picoliter well array (PWA) chip with 27,000 consistently sized picoliter reactions (314 pL) for isothermal DNA quantification using digital RPA (dRPA) at 39°C. Sample loading using a scraping liquid blade was simple, fast, and required small reagent volumes (i.e., <20 μL). Passivating the chip surface using a methoxy-PEG-silane agent effectively eliminated cross-contamination during dRPA. Our creative optical design enabled wide-field fluorescence imaging in situ and both end-point and real-time analyses of picoliter wells in a 6-cm(2) area. It was not necessary to use scan shooting and stitch serial small images together. Using this method, we quantified serial dilutions of a Listeria monocytogenes gDNA stock solution from 9 × 10(-1) to 4 × 10(-3) copies per well with an average error of less than 11% (N = 15). Overall dRPA-on-chip processing required less than 30 min, which was a 4-fold decrease compared to dPCR, requiring approximately 2 h. dRPA on the PWA chip provides a simple and highly sensitive method to quantify nucleic acids without thermal cycling or precise micropump/microvalve control. It has applications in fast field analysis and critical clinical diagnostics under resource-limited settings.

  13. Picoliter Well Array Chip-Based Digital Recombinase Polymerase Amplification for Absolute Quantification of Nucleic Acids

    PubMed Central

    Li, Zhao; Liu, Yong; Wei, Qingquan; Liu, Yuanjie; Liu, Wenwen; Zhang, Xuelian; Yu, Yude

    2016-01-01

    Absolute, precise quantification methods expand the scope of nucleic acids research and have many practical applications. Digital polymerase chain reaction (dPCR) is a powerful method for nucleic acid detection and absolute quantification. However, it requires thermal cycling and accurate temperature control, which are difficult in resource-limited conditions. Accordingly, isothermal methods, such as recombinase polymerase amplification (RPA), are more attractive. We developed a picoliter well array (PWA) chip with 27,000 consistently sized picoliter reactions (314 pL) for isothermal DNA quantification using digital RPA (dRPA) at 39°C. Sample loading using a scraping liquid blade was simple, fast, and required small reagent volumes (i.e., <20 μL). Passivating the chip surface using a methoxy-PEG-silane agent effectively eliminated cross-contamination during dRPA. Our creative optical design enabled wide-field fluorescence imaging in situ and both end-point and real-time analyses of picoliter wells in a 6-cm2 area. It was not necessary to use scan shooting and stitch serial small images together. Using this method, we quantified serial dilutions of a Listeria monocytogenes gDNA stock solution from 9 × 10-1 to 4 × 10-3 copies per well with an average error of less than 11% (N = 15). Overall dRPA-on-chip processing required less than 30 min, which was a 4-fold decrease compared to dPCR, requiring approximately 2 h. dRPA on the PWA chip provides a simple and highly sensitive method to quantify nucleic acids without thermal cycling or precise micropump/microvalve control. It has applications in fast field analysis and critical clinical diagnostics under resource-limited settings. PMID:27074005

  14. Advanced Flip Chips in Extreme Temperature Environments

    NASA Technical Reports Server (NTRS)

    Ramesham, Rajeshuni

    2010-01-01

    The use of underfill materials is necessary with flip-chip interconnect technology to redistribute stresses due to mismatching coefficients of thermal expansion (CTEs) between dissimilar materials in the overall assembly. Underfills are formulated using organic polymers and possibly inorganic filler materials. There are a few ways to apply the underfills with flip-chip technology. Traditional capillary-flow underfill materials now possess high flow speed and reduced time to cure, but they still require additional processing steps beyond the typical surface-mount technology (SMT) assembly process. Studies were conducted using underfills in a temperature range of -190 to 85 C, which resulted in an increase of reliability by one to two orders of magnitude. Thermal shock of the flip-chip test articles was designed to induce failures at the interconnect sites (-40 to 100 C). The study on the reliability of flip chips using underfills in the extreme temperature region is of significant value for space applications. This technology is considered as an enabling technology for future space missions. Flip-chip interconnect technology is an advanced electrical interconnection approach where the silicon die or chip is electrically connected, face down, to the substrate by reflowing solder bumps on area-array metallized terminals on the die to matching footprints of solder-wettable pads on the chosen substrate. This advanced flip-chip interconnect technology will significantly improve the performance of high-speed systems, productivity enhancement over manual wire bonding, self-alignment during die joining, low lead inductances, and reduced need for attachment of precious metals. The use of commercially developed no-flow fluxing underfills provides a means of reducing the processing steps employed in the traditional capillary flow methods to enhance SMT compatibility. Reliability of flip chips may be significantly increased by matching/tailoring the CTEs of the substrate

  15. Photocleavage control of nucleated DNA nanosystems--the influence of surface strand sterics.

    PubMed

    Hanna, Morcos; Munshi, Moorsalin; Kedzierski, Nancy A; Chung, Paul N; Huang, Terry; Mok, Allen K; Lukeman, Philip S

    2014-02-21

    We use sterically inaccessible 'seed' strands, released from a surface into solution by photocleavage to initiate a nucleated DNA polymerization reaction. We demonstrate control of the quantity of 'seed' release and that hairpin steric protection of the 'seed' leads to less 'leaky' surfaces. This polymerization is a model system for surface-photocleavage initiation of sub-stoichiometric reaction cascades; these cascades should find use as a component of labs-on-chips capable of bioanalytical and DNA-computing tasks. PMID:24402244

  16. Photocleavage control of nucleated DNA nanosystems--the influence of surface strand sterics.

    PubMed

    Hanna, Morcos; Munshi, Moorsalin; Kedzierski, Nancy A; Chung, Paul N; Huang, Terry; Mok, Allen K; Lukeman, Philip S

    2014-02-21

    We use sterically inaccessible 'seed' strands, released from a surface into solution by photocleavage to initiate a nucleated DNA polymerization reaction. We demonstrate control of the quantity of 'seed' release and that hairpin steric protection of the 'seed' leads to less 'leaky' surfaces. This polymerization is a model system for surface-photocleavage initiation of sub-stoichiometric reaction cascades; these cascades should find use as a component of labs-on-chips capable of bioanalytical and DNA-computing tasks.

  17. Microfluidic-integrated DNA nanobiosensors.

    PubMed

    Ansari, M I Haque; Hassan, Shabir; Qurashi, Ahsanulhaq; Khanday, Firdous Ahmad

    2016-11-15

    Over the last few decades, an increased demand has emerged for integrating biosensors with microfluidic- and nanofluidic-based lab-on-chip (LOC) devices for point-of-care (POC) diagnostics, in the medical industry and environmental monitoring of pathogenic threat agents. Such a merger of microfluidics with biosensing technologies allows for the precise control of volumes, as low as one nanolitre and the integration of various types of bioassays on a single miniaturized platform. This integration offers several favorable advantages, such as low reagent consumption, automation of sample preparation, reduction in processing time, low cost analysis, minimal handling of hazardous materials, high detection accuracy, portability and disposability. This review provides a synopsis of the most recent developments in the microfluidic-integrated biosensing field by delineating the fundamental theory of microfluidics, fabrication techniques and a detailed account of the various transduction methods that are employed. Lastly, the review discusses state-of-the-art DNA biosensors with a focus on optical DNA biosensors.

  18. Microfluidic-integrated DNA nanobiosensors.

    PubMed

    Ansari, M I Haque; Hassan, Shabir; Qurashi, Ahsanulhaq; Khanday, Firdous Ahmad

    2016-11-15

    Over the last few decades, an increased demand has emerged for integrating biosensors with microfluidic- and nanofluidic-based lab-on-chip (LOC) devices for point-of-care (POC) diagnostics, in the medical industry and environmental monitoring of pathogenic threat agents. Such a merger of microfluidics with biosensing technologies allows for the precise control of volumes, as low as one nanolitre and the integration of various types of bioassays on a single miniaturized platform. This integration offers several favorable advantages, such as low reagent consumption, automation of sample preparation, reduction in processing time, low cost analysis, minimal handling of hazardous materials, high detection accuracy, portability and disposability. This review provides a synopsis of the most recent developments in the microfluidic-integrated biosensing field by delineating the fundamental theory of microfluidics, fabrication techniques and a detailed account of the various transduction methods that are employed. Lastly, the review discusses state-of-the-art DNA biosensors with a focus on optical DNA biosensors. PMID:27179566

  19. Microchips for DNA sequencing

    NASA Astrophysics Data System (ADS)

    Mastrangelo, Carlos H.; Palaniappan, S.; Man, Piu Francis; Burns, Mark A.; Burke, David T.

    1999-08-01

    Genetic information is vital for understanding features and response of an organism. In humans, genetic errors are linked to the development of major diseases such as cancer and diabetes. In order to maximally exploit this information it is necessary to develop miniature sequencing assays that are rapid and inexpensive. In this paper we show how this could be attained with microfluidic chips that contain integrated assays. To date simple silicon/glass chips aimed for sequencing purpose have been realized; but these chips are not yet practical. Some of the solutions that are used to bring these devices closer to commercial applications are discussed.

  20. Programmable DNA-Based Finite Automata

    NASA Astrophysics Data System (ADS)

    Ratner, Tamar; Keinan, Ehud

    Computation using DNA has many advantages, including the potential for massive parallelism that allows for large number of operations per second, the direct interface between the computation process and a biological output, and the miniaturization of the computing devices to a molecular scale. In 2001, we reported on the first DNA-based, programmable finite automaton (2-symbol-2-state) capable of computing autonomously with all its hardware, software, input, and output being soluble biomolecules mixed in solution. Later, using similar principles, we developed advanced 3-symbol-3-state automata. We have also shown that real-time detection of the output signal, as well as real-time monitoring of all the computation intermediates, can be achieved by the use of surface plasmon resonance (SPR) technology. More recently, we have shown that it is possible to achieve a biologically relevant output, such as specific gene expression, by using a reporter-gene as an output-readout. We cloned the input into circular plasmids, and thereby achieved control over gene expression by a programmable sequence of computation events. Further efforts are currently directed to immobilization of the input molecules onto a solid chip to enable parallel computation, where the location of the input on the chip represents specific tagging.

  1. A versatile quantitation platform based on platinum nanoparticles incorporated volumetric bar-chart chip for highly sensitive assays.

    PubMed

    Wang, Yuzhen; Zhu, Guixian; Qi, Wenjin; Li, Ying; Song, Yujun

    2016-11-15

    Platinum nanoparticles incorporated volumetric bar-chart chip (PtNPs-V-Chip) is able to be used for point-of-care tests by providing quantitative and visualized readout without any assistance from instruments, data processing, or graphic plotting. To improve the sensitivity of PtNPs-V-Chip, hybridization chain reaction was employed in this quantitation platform for highly sensitive assays that can detect as low as 16 pM Ebola Virus DNA, 0.01ng/mL carcinoembryonic antigen (CEA), and the 10 HER2-expressing cancer cells. Based on this amplified strategy, a 100-fold decrease of detection limit was achieved for DNA by improving the number of platinum nanoparticle catalyst for the captured analyte. This quantitation platform can also distinguish single base mismatch of DNA hybridization and observe the concentration threshold of CEA. The new strategy lays the foundation for this quantitation platform to be applied in forensic analysis, biothreat detection, clinical diagnostics and drug screening.

  2. On-Chip Cellomics: Constructive Understanding of Multicellular Network Using On-Chip Cellomics Technology

    NASA Astrophysics Data System (ADS)

    Yasuda, Kenji

    2012-08-01

    We have developed methods and systems of analyzing epigenetic information in cells to expand our understanding of how living systems are determined. Because cells are minimum units reflecting epigenetic information, which is considered to map the history of a parallel-processing recurrent network of biochemical reactions, their behaviors cannot be explained by considering only conventional deonucleotide (DNA) information-processing events. The role of epigenetic information on cells, which complements their genetic information, was inferred by comparing predictions from genetic information with cell behaviour observed under conditions chosen to reveal adaptation processes and community effects. A system of analyzing epigenetic information, on-chip cellomics technology, has been developed starting from the twin complementary viewpoints of cell regulation as an “algebraic” system (emphasis on temporal aspects) and as a “geometric” system (emphasis on spatial aspects) exploiting microfabrication technology and a reconstructive approach of cellular systems not only for single cell-based subjects such as Escherichia coli and macrophages but also for cellular networks like the community effect of cardiomyocytes and plasticity in neuronal networks. One of the most important contributions of this study was to be able to reconstruct the concept of a cell regulatory network from the “local” (molecules expressed at certain times and places) to the “global” (the cell as a viable, functioning system). Knowledge of epigenetic information, which we can control and change during cell lives, complements the genetic variety, and these two types of information are indispensable for living organisms. This new knowlege has the potential to be the basis of cell-based biological and medical fields such as those involving cell-based drug screening and the regeneration of organs from stem cells.

  3. Quantitation of DNA adducts by stable isotope dilution mass spectrometry

    PubMed Central

    Tretyakova, Natalia; Goggin, Melissa; Janis, Gregory

    2012-01-01

    Exposure to endogenous and exogenous chemicals can lead to the formation of structurally modified DNA bases (DNA adducts). If not repaired, these nucleobase lesions can cause polymerase errors during DNA replication, leading to heritable mutations potentially contributing to the development of cancer. Due to their critical role in cancer initiation, DNA adducts represent mechanism-based biomarkers of carcinogen exposure, and their quantitation is particularly useful for cancer risk assessment. DNA adducts are also valuable in mechanistic studies linking tumorigenic effects of environmental and industrial carcinogens to specific electrophilic species generated from their metabolism. While multiple experimental methodologies have been developed for DNA adduct analysis in biological samples – including immunoassay, HPLC, and 32P-postlabeling – isotope dilution high performance liquid chromatography-electrospray ionization-tandem mass spectrometry (HPLC-ESI-MS/MS) generally has superior selectivity, sensitivity, accuracy, and reproducibility. As typical DNA adducts concentrations in biological samples are between 0.01 – 10 adducts per 108 normal nucleotides, ultrasensitive HPLC-ESI-MS/MS methodologies are required for their analysis. Recent developments in analytical separations and biological mass spectrometry – especially nanoflow HPLC, nanospray ionization MS, chip-MS, and high resolution MS – have pushed the limits of analytical HPLC-ESI-MS/MS methodologies for DNA adducts, allowing researchers to accurately measure their concentrations in biological samples from patients treated with DNA alkylating drugs and in populations exposed to carcinogens from urban air, drinking water, cooked food, alcohol, and cigarette smoke. PMID:22827593

  4. Utilisation of chip thickness models in grinding

    NASA Astrophysics Data System (ADS)

    Singleton, Roger

    Grinding is now a well established process utilised for both stock removal and finish applications. Although significant research is performed in this field, grinding still experiences problems with burn and high forces which can lead to poor quality components and damage to equipment. This generally occurs in grinding when the process deviates from its safe working conditions. In milling, chip thickness parameters are utilised to predict and maintain process outputs leading to improved control of the process. This thesis looks to further the knowledge of the relationship between chip thickness and the grinding process outputs to provide an increased predictive and maintenance modelling capability. Machining trials were undertaken using different chip thickness parameters to understand how these affect the process outputs. The chip thickness parameters were maintained at different grinding wheel diameters for a constant productivity process to determine the impact of chip thickness at a constant material removal rate.. Additional testing using a modified pin on disc test rig was performed to provide further information on process variables. The different chip thickness parameters provide control of different process outputs in the grinding process. These relationships can be described using contact layer theory and heat flux partitioning. The contact layer is defined as the immediate layer beneath the contact arc at the wheel workpiece interface. The size of the layer governs the force experienced during the process. The rate of contact layer removal directly impacts the net power required from the system. It was also found that the specific grinding energy of a process is more dependent on the productivity of a grinding process

  5. Expression and significance of CHIP in canine mammary gland tumors.

    PubMed

    Wang, Huanan; Yang, Xu; Jin, Yipeng; Pei, Shimin; Zhang, Di; Ma, Wen; Huang, Jian; Qiu, Hengbin; Zhang, Xinke; Jiang, Qiuyue; Sun, Weidong; Zhang, Hong; Lin, Degui

    2015-11-01

    CHIP (Carboxy terminus of Hsc70 Interacting Protein) is an E3 ubiquitin ligase that can induce ubiquitination and degradation of several oncogenic proteins. The expression of CHIP is frequently lower in human breast cancer than in normal breast tissue. However, the expression and role of CHIP in the canine mammary gland tumor (CMGT) remain unclear. We investigated the potential correlation between CHIP expression and mammary gland tumor prognosis in female dogs. CHIP expression was measured in 54 dogs by immunohistochemistry and real-time RT-PCR. CHIP protein expression was significantly correlated with the histopathological diagnosis, outcome of disease and tumor classification. The transcriptional level of CHIP was significantly higher in normal tissues (P=0.001) and benign tumors (P=0.009) than it in malignant tumors. CHIP protein expression was significantly correlated with the transcriptional level of CHIP (P=0.0102). The log-rank test survival curves indicated that patients with low expression of CHIP had shorter overall periods of survival than those with higher CHIP protein expression (P=0.050). Our data suggest that CHIP may play an important role in the formation and development of CMGTs and serve as a valuable prognostic marker and potential target for genetic therapy.

  6. 7. VIEW OF THE CHIP ROASTER LOCATED IN BUILDING 447. ...

    Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

    7. VIEW OF THE CHIP ROASTER LOCATED IN BUILDING 447. THE CHIP ROASTER BURNED URANIUM CHIPS FROM MACHINING AREAS TO AN OXIDE, A MORE STABLE FORM FOR DISPOSAL. (4/27/55) - Rocky Flats Plant, Non-Nuclear Production Facility, South of Cottonwood Avenue, west of Seventh Avenue & east of Building 460, Golden, Jefferson County, CO

  7. Integrated Carbon Nanotubes Electrodes in Microfluidic Chip via MWPCVD

    NASA Astrophysics Data System (ADS)

    Wang, Shenggao; Wang, Mingyang; Yu, Dongdong; Zhang, Wenbo; Deng, Xiaoqing; Du, Yu; Cheng, Lili; Wang, Jianhua

    2010-10-01

    An on-chip electrochemical detector for microfluidic chips was described, based on integrated carbon nanotube (CNT) electrodes directly onto the chip substrate through microwave plasma chemical vapor deposition (MWPCVD). The attractive performance of the integrated CNT electrodes was demonstrated for the amperometric detection of sucrose, glucose and D-fructose. The integrated CNT electrodes showed stronger electrocatalytic activity than gold electrodes.

  8. Chip forwarder for mobile in-woods chippers

    SciTech Connect

    Duncan, M.R.; Gibson, H.G.; Krutz, G.W.; Pope, P.E.

    1984-01-01

    A forwarder-based prehauler to service a mobile, wholetree chip harvester was designed. The vacuum pneumatic conveyor uses airflow to load and unload chips, chips being fed into the airflow by rotating pipes with feed scoops and internal augers. Initial tests measured main parameters and confirmed concept feasibility. 12 references.

  9. Smart single-chip gas sensor microsystem.

    PubMed

    Hagleitner, C; Hierlemann, A; Lange, D; Kummer, A; Kerness, N; Brand, O; Baltes, H

    2001-11-15

    Research activity in chemical gas sensing is currently directed towards the search for highly selective (bio)chemical layer materials, and to the design of arrays consisting of different partially selective sensors that permit subsequent pattern recognition and multi-component analysis. Simultaneous use of various transduction platforms has been demonstrated, and the rapid development of integrated-circuit technology has facilitated the fabrication of planar chemical sensors and sensors based on three-dimensional microelectromechanical systems. Complementary metal-oxide silicon processes have previously been used to develop gas sensors based on metal oxides and acoustic-wave-based sensor devices. Here we combine several of these developments to fabricate a smart single-chip chemical microsensor system that incorporates three different transducers (mass-sensitive, capacitive and calorimetric), all of which rely on sensitive polymeric layers to detect airborne volatile organic compounds. Full integration of the microelectronic and micromechanical components on one chip permits control and monitoring of the sensor functions, and enables on-chip signal amplification and conditioning that notably improves the overall sensor performance. The circuitry also includes analog-to-digital converters, and an on-chip interface to transmit the data to off-chip recording units. We expect that our approach will provide a basis for the further development and optimization of gas microsystems.

  10. Protein-DNA binding in high-resolution

    PubMed Central

    Mahony, Shaun; Pugh, B. Franklin

    2015-01-01

    Recent advances in experimental and computational methodologies are enabling ultra-high resolution genome-wide profiles of protein-DNA binding events. For example, the ChIP-exo protocol precisely characterizes protein-DNA crosslinking patterns by combining chromatin immunoprecipitation (ChIP) with 5′ → 3′ exonuclease digestion. Similarly, deeply sequenced chromatin accessibility assays (e.g. DNase-seq and ATACseq) enable the detection of protected footprints at protein-DNA binding sites. With these techniques and others, we have the potential to characterize the individual nucleotides that interact with transcription factors, nucleosomes, RNA polymerases, and other regulatory proteins in a particular cellular context. In this review, we explain the experimental assays and computational analysis methods that enable high-resolution profiling of protein-DNA binding events. We discuss the challenges and opportunities associated with such approaches. PMID:26038153

  11. Reliability evaluation of CIF (chip-in-flex) and COF (chip-on-flex) packages

    NASA Astrophysics Data System (ADS)

    Jang, Jae-Won; Suk, Kyoung-Lim; Paik, Kyung-Wook; Lee, Soon-Bok

    2009-12-01

    CIF (chip-in-flex) and COF (chip-on-flex) packages have the advantages of fine pitch capability, and flexibility. Anisotropic conductive films (ACFs) are used for the interconnection between chip and substrate. Display, mobile device, and semiconductor industry require for smaller and more integrated packages. Both CIF and COF packages are an alternative for the demands. However, there are some reliability problems of interconnection between the chip and substrate because the packages are subjected to various loading conditions. These may degrade the functionality of the packages. Therefore, reliability assessment of both packages is necessary. In this study, experimental tests were performed to evaluate the reliability of interconnection between the chip and substrate of CIF and COF packages. Thermal cycling tests were performed to evaluate the resistance against thermal fatigue. The shape and warpage of the chip of CIF and COF packages were observed using optical methods (e.g., shadow Moiré and Twyman/Green interferometry). These optical Moiré techniques are widely used for measuring small deformations in microelectronic packages. The stress distribution around the chip was evaluated through FEA (finite element analysis). In addition, we suggested modifying design parameter of CIF packages for the reliability enhancement.

  12. Reliability evaluation of CIF (chip-in-flex) and COF (chip-on-flex) packages

    NASA Astrophysics Data System (ADS)

    Jang, Jae-Won; Suk, Kyoung-Lim; Paik, Kyung-Wook; Lee, Soon-Bok

    2010-03-01

    CIF (chip-in-flex) and COF (chip-on-flex) packages have the advantages of fine pitch capability, and flexibility. Anisotropic conductive films (ACFs) are used for the interconnection between chip and substrate. Display, mobile device, and semiconductor industry require for smaller and more integrated packages. Both CIF and COF packages are an alternative for the demands. However, there are some reliability problems of interconnection between the chip and substrate because the packages are subjected to various loading conditions. These may degrade the functionality of the packages. Therefore, reliability assessment of both packages is necessary. In this study, experimental tests were performed to evaluate the reliability of interconnection between the chip and substrate of CIF and COF packages. Thermal cycling tests were performed to evaluate the resistance against thermal fatigue. The shape and warpage of the chip of CIF and COF packages were observed using optical methods (e.g., shadow Moiré and Twyman/Green interferometry). These optical Moiré techniques are widely used for measuring small deformations in microelectronic packages. The stress distribution around the chip was evaluated through FEA (finite element analysis). In addition, we suggested modifying design parameter of CIF packages for the reliability enhancement.

  13. A Low-Cost Microfluidic Chip for Rapid Genotyping of Malaria-Transmitting Mosquitoes

    PubMed Central

    Liu, Changchun; Mauk, Michael G.; Hart, Robert; Bonizzoni, Mariangela; Yan, Guiyun; Bau, Haim H.

    2012-01-01

    Background Vector control is one of the most effective measures to prevent the transmission of malaria, a disease that causes over 600,000 deaths annually. Around 30–40 Anopheles mosquito species are natural vectors of malaria parasites. Some of these species cannot be morphologically distinguished, but have behavioral and ecological differences. Emblematic of this is the Anopheles gambiae species complex. The correct identification of vector species is fundamental to the development of control strategies and epidemiological studies of disease transmission. Methodology/Principal Findings An inexpensive, disposable, field-deployable, sample-to-answer, microfluidic chip was designed, constructed, and tested for rapid molecular identification of Anopheles gambiae and Anopheles arabiensis. The chip contains three isothermal amplification reactors. One test reactor operates with specific primers to amplify Anopheles gambiae DNA, another with specific primers for Anopheles arabiensis DNA, and the third serves as a negative control. A mosquito leg was crushed on an isolation membrane. Two discs, laden with mosquito tissue, were punched out of the membrane and inserted into the two test chambers. The isolated, disc-bound DNA served as a template in the amplification processes. The amplification products were detected with intercalating fluorescent dye that was excited with a blue light-emitting diode. The emitted light was observed by eye and recorded with a cell-phone camera. When the target consisted of Anopheles gambiae, the reactor containing primers specific to An. gambiae lit up while the other two reactors remained dark. When the target consisted of Anopheles arabiensis, the reactor containing primers specific to An. arabiensis lit up while the other two reactors remained dark. Conclusions/Significance The microfluidic chip provides a means to identify mosquito type through molecular analysis. It is suitable for field work, allowing one to track the geographical

  14. A tripartite paternally methylated region within the Gpr1-Zdbf2 imprinted domain on mouse chromosome 1 identified by meDIP-on-chip

    PubMed Central

    Hiura, Hitoshi; Sugawara, Atsushi; Ogawa, Hidehiko; John, Rosalind M.; Miyauchi, Naoko; Miyanari, Yusuke; Horiike, Tokumasa; Li, Yufeng; Yaegashi, Nobuo; Sasaki, Hiroyuki; Kono, Tomohiro; Arima, Takahiro

    2010-01-01

    The parent-of-origin specific expression of imprinted genes relies on DNA methylation of CpG-dinucleotides at differentially methylated regions (DMRs) during gametogenesis. To date, four paternally methylated DMRs have been identified in screens based on conventional approaches. These DMRs are linked to the imprinted genes H19, Gtl2 (IG-DMR), Rasgrf1 and, most recently, Zdbf2 which encodes zinc finger, DBF-type containing 2. In this study, we applied a novel methylated-DNA immunoprecipitation-on-chip (meDIP-on-chip) method to genomic DNA from mouse parthenogenetic- and androgenetic-derived stem cells and sperm and identified 458 putative DMRs. This included the majority of known DMRs. We further characterized the paternally methylated Zdbf2/ZDBF2 DMR. In mice, this extensive germ line DMR spanned 16 kb and possessed an unusual tripartite structure. Methylation was dependent on DNA methyltransferase 3a (Dnmt3a), similar to H19 DMR and IG-DMR. In both humans and mice, the adjacent gene, Gpr1/GPR1, which encodes a G-protein-coupled receptor 1 protein with transmembrane domain, was also imprinted and paternally expressed. The Gpr1-Zdbf2 domain was most similar to the Rasgrf1 domain as both DNA methylation and the actively expressed allele were in cis on the paternal chromosome. This work demonstrates the effectiveness of meDIP-on-chip as a technique for identifying DMRs. PMID:20385583

  15. Detection of tumor markers with ProteinChip technology.

    PubMed

    Wiesner, Andreas

    2004-02-01

    The early diagnosis of cancer at a curable stage is crucial for the successful treatment of this disease. Most of the currently used tumor assays appear too late and rely on single biomarkers with high false-negative and/or false-positive rates. As an additional burden for the patient, the traditional assays often require biopsy material instead of less invasively taken samples like serum. With the hope for more reliable DNA- and RNA-based screening tools, the research activities of the past 20 years have focused on the genomic characteristics of cancer cells. But, up to now, the output from this strategy has been disappointingly low and the disillusionment is paired with a return to proteins as the real key players in all physiological and pathological processes. Meanwhile, comparative protein profiling is generally acknowledged as a promising way for the detection of specific and predictive protein patterns reflecting certain stages of cancer without dependency on single markers. To meet the new technological demands, the ProteinChip Biomarker System was developed for the Expression Difference Mapping analysis of several hundreds of samples per day on a single, uncomplicated platform; with software support for the construction of multi-marker predictive models. The Interaction Discovery Mapping platform is introduced as the next methodical step for investigations about protein binding partners of possible importance in diagnosis and therapy. This review summarizes the current state in cancer diagnosis, provides an introduction into the ProteinChip technology, and gives an update on publications and research collaborations in SELDI-based tumor marker discovery.

  16. Optical DNA

    NASA Astrophysics Data System (ADS)

    Vijaywargi, Deepak; Lewis, Dave; Kirovski, Darko

    A certificate of authenticity (COA) is an inexpensive physical object with a random and unique structure S which is hard to near-exactly replicate. An inexpensive device should be able to scan object’s physical “fingerprint,” a set of features that represents S. In this paper, we explore one set of requirements that optical media such as DVDs should satisfy, to be considered as COAs. As manufacturing of such media produces inevitable errors, we use the locations and count of these errors as a “fingerprint” for each optical disc: its optical DNA. The “fingerprint” is signed using publisher’s private-key and the resulting signature is stored onto the optical medium using a post-production process. Standard DVD players with altered firmware that includes publisher’s public-key, should be able to verify the authenticity of DVDs protected with optical DNA. Our key finding is that for the proposed protocol, only DVDs with exceptional wear-and-tear characteristics would result in an inexpensive and viable anti-counterfeiting technology.

  17. CHIPS Neutrino Detector Research and Development

    NASA Astrophysics Data System (ADS)

    Salazar, Ramon; Vahle, Patricia; Chips Collaboration

    2015-04-01

    The CHIPS R&D project is an effort to develop affordable megaton-scale neutrino detectors. The CHIPS strategy calls for submerging water Cherenkov detectors deep under water. The surrounding water acts as structural support, minimizing large initial investments in costly infrastructure, and serves as an overburden, shielding the detector from cosmic rays and eliminating the need for expensive underground construction. Additional cost savings will be achieved through photodetector development and optimization of readout geometry. In summer 2014 a small prototype of the CHIPS detector was deployed in the flooded Wentworth Mine Pit in Northern Minnesota. The detector has been recording data underwater throughout the fall and winter. In this talk, we will discuss lessons learned from the prototyping experience and the plans for submerging much larger detectors in future years.

  18. A compact PE memory for vision chips

    NASA Astrophysics Data System (ADS)

    Cong, Shi; Zhe, Chen; Jie, Yang; Nanjian, Wu; Zhihua, Wang

    2014-09-01

    This paper presents a novel compact memory in the processing element (PE) for single-instruction multiple-data (SIMD) vision chips. The PE memory is constructed with 8 × 8 register cells, where one latch in the slave stage is shared by eight latches in the master stage. The memory supports simultaneous read and write on the same address in one clock cycle. Its compact area of 14.33 μm2/bit promises a higher integration level of the processor. A prototype chip with a 64 × 64 PE array is fabricated in a UMC 0.18 μm CMOS technology. Five types of the PE memory cell structure are designed and compared. The testing results demonstrate that the proposed PE memory architecture well satisfies the requirement of the vision chip in high-speed real-time vision applications, such as 1000 fps edge extraction.

  19. Time of flight system on a chip

    NASA Technical Reports Server (NTRS)

    Paschalidis, Nicholas P. (Inventor)

    2006-01-01

    A CMOS time-of-flight TOF system-on-a-chip SoC for precise time interval measurement with low power consumption and high counting rate has been developed. The analog and digital TOF chip may include two Constant Fraction Discriminators CFDs and a Time-to-Digital Converter TDC. The CFDs can interface to start and stop anodes through two preamplifiers and perform signal processing for time walk compensation (110). The TDC digitizes the time difference with reference to an off-chip precise external clock (114). One TOF output is an 11-bit digital word and a valid event trigger output indicating a valid event on the 11-bit output bus (116).

  20. Design of highway embankments using tire chips

    SciTech Connect

    Bosscher, P.J.; Edil, T.B.; Kuraoka, S.

    1997-04-01

    This paper describes research undertaken to develop design procedures for using shredded scrap tires as a lightweight fill material in highway construction. The benefits of using scrap tires are particularly enhanced if they can be used to replace virgin construction materials made from nonrenewable resources. This paper addresses the use of tire chips as a highway embankment material. Design parameters for embankments constructed using discarded shredded tires are presented based on laboratory model studies, numerical analyses, and field performance of test fills. The conclusions of this report support the use of tire chips as an environmentally acceptable lightweight fill in highway applications if properly confined. Recommendations for design procedures and construction specifications for the use of tire chips in highway fills are provided.

  1. A proposed holistic approach to on-chip, off-chip, test, and package interconnections

    NASA Astrophysics Data System (ADS)

    Bartelink, Dirk J.

    1998-11-01

    The term interconnection has traditionally implied a `robust' connection from a transistor or a group of transistors in an IC to the outside world, usually a PC board. Optimum system utilization is done from outside the IC. As an alternative, this paper addresses `unimpeded' transistor-to-transistor interconnection aimed at reaching the high circuit densities and computational capabilities of neighboring IC's. In this view, interconnections are not made to some human-centric place outside the IC world requiring robustness—except for system input and output connections. This unimpeded interconnect style is currently available only through intra-chip signal traces in `system-on-a-chip' implementations, as exemplified by embedded DRAMs. Because the traditional off-chip penalty in performance and wiring density is so large, a merging of complex process technologies is the only option today. It is suggested that, for system integration to move forward, the traditional robustness requirement inherited from conventional packaging interconnect and IC manufacturing test must be discarded. Traditional system assembly from vendor parts requires robustness under shipping, inspection and assembly. The trend toward systems on a chip signifies willingness by semiconductor companies to design and fabricate whole systems in house, so that `in-house' chip-to-chip assembly is not beyond reach. In this scenario, bare chips never leave the controlled environment of the IC fabricator while the two major contributors to off-chip signal penalty, ESD protection and the need to source a 50-ohm test head, are avoided. With in-house assembly, ESD protection can be eliminated with the precautions already familiar in plasma etching. Test interconnection impacts the fundamentals of IC manufacturing, particularly with clock speeds approaching 1GHz, and cannot be an afterthought. It should be an integral part of the chip-to-chip interconnection bandwidth optimization, because—as we must

  2. Biostability of an implantable glucose sensor chip

    NASA Astrophysics Data System (ADS)

    Fröhlich, M.; Birkholz, M.; Ehwald, K. E.; Kulse, P.; Fursenko, O.; Katzer, J.

    2012-12-01

    Surface materials of an implantable microelectronic chip intended for medical applications were evaluated with respect to their long-term stability in bio-environments. The sensor chip shall apply in a glucose monitor by operating as a microviscosimeter according to the principle of affinity viscosimetry. A monolithic integration of a microelectromechanical system (MEMS) into the sensor chip was successfully performed in a combined 0.25 μm CMOS/BiCMOS technology. In order to study material durability and biostability of the surfaces, sensor chips were exposed to various in vitro and in vivo tests. Corrosional damage of SiON, SiO2 and TiN surfaces was investigated by optical microscopy, ellipsometry and AFM. The results served for optimizing the Back-end-of-Line (BEoL) stack, from which the MEMS was prepared. Corrosion of metal lines could significantly be reduced by improving the topmost passivation layer. The experiments revealed no visible damage of the actuator or other functionally important MEMS elements. Sensor chips were also exposed to human body fluid for three month by implantation into the abdomen of a volunteer. Only small effects were observed for layer thickness and Ra roughness after explantation. In particular, TiN as used for the actuator beam showed no degradation by biocorrosion. The highest degradation rate of about 50 nm per month was revealed for the SiON passivation layer. These results suggest that the sensor chip may safely operate in subcutaneous tissue for a period of several months.

  3. Lab-on a-Chip

    NASA Technical Reports Server (NTRS)

    1999-01-01

    Labs on chips are manufactured in many shapes and sizes and can be used for numerous applications, from medical tests to water quality monitoring to detecting the signatures of life on other planets. The eight holes on this chip are actually ports that can be filled with fluids or chemicals. Tiny valves control the chemical processes by mixing fluids that move in the tiny channels that look like lines, connecting the ports. Scientists at NASA's Marshall Space Flight Center (MSFC) in Huntsville, Alabama designed this chip to grow biological crystals on the International Space Station (ISS). Through this research, they discovered that this technology is ideally suited for solving the challenges of the Vision for Space Exploration. For example, thousands of chips the size of dimes could be loaded on a Martian rover looking for biosignatures of past or present life. Other types of chips could be placed in handheld devices used to monitor microbes in water or to quickly conduct medical tests on astronauts. The portable, handheld Lab-on-a Chip Application Development Portable Test System (LOCAD-PTS) made its debut flight aboard Discovery during the STS-116 mission launched December 9, 2006. The system allowed crew members to monitor their environment for problematic contaminants such as yeast, mold, and even E.coli, and salmonella. Once LOCAD-PTS reached the ISS, the Marshall team continued to manage the experiment, monitoring the study from a console in the Payload Operations Center at MSFC. The results of these studies will help NASA researchers refine the technology for future Moon and Mars missions. (NASA/MSFC/D.Stoffer)

  4. CHIP: A new modulator of human malignant disorders

    PubMed Central

    Shao, Qianqian; Yang, Gang; Zheng, Lianfang; Zhang, Taiping; Zhao, Yupei

    2016-01-01

    Carboxyl terminus of Hsc70-interacting protein (CHIP) is known as a chaperone-associated E3 for a variety of protein substrates. It acts as a link between molecular chaperones and ubiquitin–proteasome system. Involved in the process of protein clearance, CHIP plays a critical role in maintaining protein homeostasis in diverse conditions. Here, we provide a comprehensive review of our current understanding of CHIP and summarize recent advances in CHIP biology, with a focus on CHIP in the setting of malignancies. PMID:27007160

  5. CHIP: A new modulator of human malignant disorders.

    PubMed

    Cao, Zhe; Li, Guanqiao; Shao, Qianqian; Yang, Gang; Zheng, Lianfang; Zhang, Taiping; Zhao, Yupei

    2016-05-17

    Carboxyl terminus of Hsc70-interacting protein (CHIP) is known as a chaperone-associated E3 for a variety of protein substrates. It acts as a link between molecular chaperones and ubiquitin-proteasome system. Involved in the process of protein clearance, CHIP plays a critical role in maintaining protein homeostasis in diverse conditions. Here, we provide a comprehensive review of our current understanding of CHIP and summarize recent advances in CHIP biology, with a focus on CHIP in the setting of malignancies.

  6. Standardization of Spore Inactivation Method for PMA-PhyloChip Analysis

    NASA Technical Reports Server (NTRS)

    Schrader, Michael

    2011-01-01

    In compliance with the Committee on Space Research (COSPAR) planetary protection policy, National Aeronautics and Space Administration (NASA) monitors the total microbial burden of spacecraft as a means for minimizing the inadvertent transfer of viable contaminant microorganisms to extraterrestrial environments (forward contamination). NASA standard assay-based counts are used both as a proxy for relative surface cleanliness and to estimate overall microbial burden as well as to assess whether forward planetary protection risk criteria are met for a given mission, which vary by the planetary body to be explored and whether or not life detection missions are present. Despite efforts to reduce presence of microorganisms from spacecraft prior to launch, microbes have been isolated from spacecraft and associated surfaces within the extreme conditions of clean room facilities using state of the art molecular technologies. Development of a more sensitive method that will better enumerate all viable microorganisms from spacecraft and associated surfaces could support future life detection missions. Current culture-based (NASA standard spore assay) and nucleic-acid-based polymerase chain reaction (PCR) methods have significant shortcomings in this type of analysis. The overall goal of this project is to evaluate and validate a new molecular method based on the use of a deoxyribonucleic acid (DNA) intercalating agent propidium monoazide (PMA). This is used in combination with DNA microarray (PhyloChip) which has been shown to identify very low levels of organisms on spacecraft associated surfaces. PMA can only penetrate the membrane of dead cells. Once penetrated, it intercalates the DNA and, upon photolysis using visible light it produces stable DNA monoadducts. This allows DNA to be unavailable for further PCR analysis. The specific aim of this study is to standardize the spore inactivation method for PMA-PhyloChip analysis. We have used the bacterial spores Bacillus

  7. CRRES microelectronic test chip orbital data. II

    NASA Technical Reports Server (NTRS)

    Soli, G. A.; Blaes, B. R.; Buehler, M. G.; Ray, K.; Lin, Y.-S.

    1992-01-01

    Data from a MOSFET matrix on two JPL (CIT Jet Propulsion Laboratory) CRRES (Combined Release and Radiation Effects Satellite) chips, each behind different amounts of shielding, are presented. Space damage factors are nearly identical to ground test values for pMOSFETs. The results from neighboring rows of MOSFETs show similar radiation degradation. The SRD (Space Radiation Dosimeter) is used to measure the total dose accumulated by the JPL chips. A parameter extraction algorithm that does not underestimate threshold voltage shifts is used. Temperature effects are removed from the MOSFET data.

  8. Miniature integrated-optical wavelength analyzer chip

    NASA Astrophysics Data System (ADS)

    Kunz, R. E.; Dübendorfer, J.

    1995-11-01

    A novel integrated-optical chip suitable for realizing compact miniature wavelength analyzers with high linear dispersion is presented. The chip performs the complete task of converting the spectrum of an input beam into a corresponding spatial irradiance distribution without the need for an imaging function. We demonstrate the feasibility of this approach experimentally by monitoring the changes in the mode spectrum of a laser diode on varying its case temperature. Comparing the results with simultaneous measurements by a commercial spectrometer yielded a rms wavelength deviation of 0.01 nm.

  9. Realization of a Superconducting Atom Chip

    SciTech Connect

    Nirrengarten, T.; Qarry, A.; Roux, C.; Emmert, A.; Nogues, G.; Brune, M.; Raimond, J.-M.; Haroche, S.

    2006-11-17

    We have trapped rubidium atoms in the magnetic field produced by a superconducting atom chip operated at liquid helium temperatures. Up to 8.2x10{sup 5} atoms are held in a Ioffe-Pritchard trap at a distance of 440 {mu}m from the chip surface, with a temperature of 40 {mu}K. The trap lifetime reaches 115 s at low atomic densities. These results open the way to the exploration of atom-surface interactions and coherent atomic transport in a superconducting environment, whose properties are radically different from normal metals at room temperature.

  10. MCMII and the TriP chip

    SciTech Connect

    Juan Estrada et al.

    2003-12-19

    We describe the development of the electronics that will be used to read out the Fiber Tracker and Preshower detectors in Run IIb. This electronics is needed for operation at 132ns bunch crossing, and may provide a measurement of the z coordinate of the Fiber Tracker hits when operating at 396ns bunch crossing. Specifically, we describe the design and preliminary tests of the Trip chip, MCM IIa, MCM IIb and MCM IIc. This document also serves as a user manual for the Trip chip and the MCM.

  11. Laser wavelength metrology with color sensor chips.

    PubMed

    Jones, Tyler B; Otterstrom, Nils; Jackson, Jarom; Archibald, James; Durfee, Dallin S

    2015-12-14

    We present a laser wavelength meter based on a commercial color sensor chip. The chip consists of an array of photodiodes with different absorptive color filters. By comparing the relative amplitudes of light on the photodiodes, the wavelength of light can be determined. In addition to absorption in the filters, etalon effects add additional spectral features which improve the precision of the device. Comparing the measurements from the device to a commercial wavelength meter and to an atomic reference, we found that the device has picometer-level precision and picometer-scale drift over a period longer than a month. PMID:26699036

  12. Novel PbS detector chip pattern with extinction function

    NASA Astrophysics Data System (ADS)

    Chen, Fengjin; Si, Junjie; Su, Xianjun; Lv, Yanqiu; Shi, Zhengfeng

    2015-10-01

    A novel chip pattern with extinction function in Lead salt detectors is specified. Lead Sulfide (PbS) polycrystalline film is prepared by Chemical Bath Deposition (CMD) on a transparent substrate, then a special figure and structure is saved by lithography techonology on the substrate. As a quaternion detector chip that made by PbS thin film for example in this paper, whose performance including signal, noise, weak-peaks and the uniformity of the chip are too poor to meet the detecting system at the initial stage of research, and the qualified ratio of chips is only 3% .This paper explains the reason why the performance and qualified ratio of chips were so poor, focuses on a novel chip pattern with extinction which avoided the disadvantages of traditional one. the novel chip pattern has been applied in detectors. The novel chip pattern is prepared with PbS thin film which both "extinction slice" and detector chip are based on a same substrate , which not only had absorbed the jumbled light , improved the uniformity and other performance of photosensitive elements, but also had left out the assembly diffculty and precision demand when a extinction slice assembly in the restricted space of inswept detector chip, omitted the production process of extinction slice and shorten the assembly process of the detectors, and the qualified ratio of chips had been improved from 3% to 98%.

  13. On-chip quantitative detection of pathogen genes by autonomous microfluidic PCR platform.

    PubMed

    Tachibana, Hiroaki; Saito, Masato; Shibuya, Shogo; Tsuji, Koji; Miyagawa, Nobuyuki; Yamanaka, Keiichiro; Tamiya, Eiichi

    2015-12-15

    Polymerase chain reaction (PCR)-based genetic testing has become a routine part of clinical diagnoses and food testing. In these fields, rapid, easy-to-use, and cost-efficient PCR chips are expected to be appeared for providing such testing on-site. In this study, a new autonomous disposable plastic microfluidic PCR chip was created, and was utilized for quantitative detection of pathogenic microorganisms. To control the capillary flow of the following solution in the PCR microchannel, a driving microchannel was newly designed behind the PCR microchannel. This allowed the effective PCR by simply dropping the PCR solution onto the inlet without any external pumps. In order to achieve disposability, injection-molded cyclo-olefin polymer (COP) of a cost-competitive plastic was used for the PCR chip. We discovered that coating the microchannel walls with non-ionic surfactant produced a suitable hydrophilic surface for driving the capillary flow through the 1250-mm long microchannel. As a result, quantitative real-time PCR with the lowest initial concentration of human, Escherichia coli (E. coli), and pathogenic E. coli O157 genomic DNA of 4, 0.0019, 0.031 pg/μl, respectively, was successfully achieved in less than 18 min. Our results indicate that the platform presented in this study provided a rapid, easy-to-use, and low-cost real-time PCR system that could be potentially used for on-site gene testing.

  14. The AmpliChip CYP450 test: principles, challenges, and future clinical utility in digestive disease.

    PubMed

    Juran, Brian D; Egan, Laurence J; Lazaridis, Konstantinos N

    2006-07-01

    Understanding genetically encoded inherited differences in drug metabolism and targets (ie, receptors, transporters) offers the promise of minimizing adverse drug reactions and improving therapies. Among the enzymes involved in drug metabolism, the cytochromes P450 (CYP450) hold a central position. In fact, CYP450 are involved in the biotransformation of most drugs used in clinical practice. Recent advances in the development of DNA-based diagnostics, coupled with a better understanding of genetic polymorphisms in influencing pharmacologic responses, have provided the foundation for novel in vitro tests that may predict side effects and/or therapeutic responses. The AmpliChip CYP450 test was developed as a clinical test to evaluate an individual's metabolic capacity for certain drugs by identifying polymorphisms of 2 CYP450 enzymes (ie, CYP2D6 and CYP2D19). Even though the AmpliChip CYP450 has been approved by the US Food and Drug Administration, its practical clinical utility has not yet been determined, and there is a paucity of data related to gastrointestinal and liver diseases. An understanding of the principles and opportunities provided by this new category of diagnostic test is key before planning the necessary studies to evaluate the usefulness of AmpliChip CYP450 in gastroenterologic clinical practice.

  15. GeneChip{sup {trademark}} screening assay for cystic fibrosis mutations

    SciTech Connect

    Cronn, M.T.; Miyada, C.G.; Fucini, R.V.

    1994-09-01

    GeneChip{sup {trademark}} assays are based on high density, carefully designed arrays of short oligonucleotide probes (13-16 bases) built directly on derivatized silica substrates. DNA target sequence analysis is achieved by hybridizing fluorescently labeled amplification products to these arrays. Fluorescent hybridization signals located within the probe array are translated into target sequence information using the known probe sequence at each array feature. The mutation screening assay for cystic fibrosis includes sets of oligonucleotide probes designed to detect numerous different mutations that have been described in 14 exons and one intron of the CFTR gene. Each mutation site is addressed by a sub-array of at least 40 probe sequences, half designed to detect the wild type gene sequence and half designed to detect the reported mutant sequence. Hybridization with homozygous mutant, homozygous wild type or heterozygous targets results in distinctive hybridization patterns within a sub-array, permitting specific discrimination of each mutation. The GeneChip probe arrays are very small (approximately 1 cm{sup 2}). There miniature size coupled with their high information content make GeneChip probe arrays a useful and practical means for providing CF mutation analysis in a clinical setting.

  16. Binding of leachable components of polymethyl methacrylate (PMMA) and peptide on modified SPR chip

    NASA Astrophysics Data System (ADS)

    Szaloki, M.; Vitalyos, G.; Harfalvi, J.; Hegedus, Cs

    2013-12-01

    Many types of polymers are often used in dentistry, which may cause allergic reaction, mainly methyl methacrylate allergy due to the leachable, degradable components of polymerized dental products. The aim of this study was to investigate the interaction between the leachable components of PMMA and peptides by Fourier-transform Surface Plasmon Resonance (FT SPR). In our previous work binding of oligopeptides (Ph.D.-7 and Ph.D.-12 Peptide Library Kit) was investigated to PMMA surface by phage display technique. It was found that oligopeptides bounded specifically to PMMA surface. The most common amino acids were leucine and proline inside the amino acids sequences of DNA of phages. The binding of haptens, as formaldehyde and methacrylic acid, to frequent amino acids was to investigate on the modified gold SPR chip. Self assembled monolayer (SAM) modified the surface of gold chip and ensured the specific binding between the haptens and amino acids. It was found that amino acids bounded to modified SPR gold and the haptens bounded to amino acids by creating multilayer on the chip surface. By the application of phage display and SPR modern bioanalytical methods the interaction between allergens and peptides can be investigated.

  17. Spotting and validation of a genome wide oligonucleotide chip with duplicate measurement of each gene

    SciTech Connect

    Thomassen, Mads . E-mail: mads.thomassen@ouh.fyns-amt.dk; Skov, Vibe; Eiriksdottir, Freyja; Tan, Qihua; Jochumsen, Kirsten; Fritzner, Niels; Brusgaard, Klaus; Dahlgaard, Jesper; Kruse, Torben A.

    2006-06-16

    The quality of DNA microarray based gene expression data relies on the reproducibility of several steps in a microarray experiment. We have developed a spotted genome wide microarray chip with oligonucleotides printed in duplicate in order to minimise undesirable biases, thereby optimising detection of true differential expression. The validation study design consisted of an assessment of the microarray chip performance using the MessageAmp and FairPlay labelling kits. Intraclass correlation coefficient (ICC) was used to demonstrate that MessageAmp was significantly more reproducible than FairPlay. Further examinations with MessageAmp revealed the applicability of the system. The linear range of the chips was three orders of magnitude, the precision was high, as 95% of measurements deviated less than 1.24-fold from the expected value, and the coefficient of variation for relative expression was 13.6%. Relative quantitation was more reproducible than absolute quantitation and substantial reduction of variance was attained with duplicate spotting. An analysis of variance (ANOVA) demonstrated no significant day-to-day variation.

  18. Radiolabelling diverse positron emission tomography (PET) tracers using a single digital microfluidic reactor chip.

    PubMed

    Chen, Supin; Javed, Muhammad Rashed; Kim, Hee-Kwon; Lei, Jack; Lazari, Mark; Shah, Gaurav J; van Dam, R Michael; Keng, Pei-Yuin; Kim, Chang-Jin C J

    2014-03-01

    Radiotracer synthesis is an ideal application for microfluidics because only nanogram quantities are needed for positron emission tomography (PET) imaging. Thousands of radiotracers have been developed in research settings but only a few are readily available, severely limiting the biological problems that can be studied in vivo via PET. We report the development of an electrowetting-on-dielectric (EWOD) digital microfluidic chip that can synthesize a variety of (18)F-labeled tracers targeting a range of biological processes by confirming complete syntheses of four radiotracers: a sugar, a DNA nucleoside, a protein labelling compound, and a neurotransmitter. The chip employs concentric multifunctional electrodes that are used for heating, temperature sensing, and EWOD actuation. All of the key synthesis steps for each of the four (18)F-labeled tracers are demonstrated and characterized with the chip: concentration of fluoride ion, solvent exchange, and chemical reactions. The obtained fluorination efficiencies of 90-95% are comparable to, or greater than, those achieved by conventional approaches.

  19. On-chip real-time single-copy polymerase chain reaction in picoliter droplets

    SciTech Connect

    Beer, N R; Hindson, B; Wheeler, E; Hall, S B; Rose, K A; Kennedy, I; Colston, B

    2007-04-20

    The first lab-on-chip system for picoliter droplet generation and PCR amplification with real-time fluorescence detection has performed PCR in isolated droplets at volumes 10{sup 6} smaller than commercial real-time PCR systems. The system utilized a shearing T-junction in a silicon device to generate a stream of monodisperse picoliter droplets that were isolated from the microfluidic channel walls and each other by the oil phase carrier. An off-chip valving system stopped the droplets on-chip, allowing them to be thermal cycled through the PCR protocol without droplet motion. With this system a 10-pL droplet, encapsulating less than one copy of viral genomic DNA through Poisson statistics, showed real-time PCR amplification curves with a cycle threshold of {approx}18, twenty cycles earlier than commercial instruments. This combination of the established real-time PCR assay with digital microfluidics is ideal for isolating single-copy nucleic acids in a complex environment.

  20. On-chip quantitative detection of pathogen genes by autonomous microfluidic PCR platform.

    PubMed

    Tachibana, Hiroaki; Saito, Masato; Shibuya, Shogo; Tsuji, Koji; Miyagawa, Nobuyuki; Yamanaka, Keiichiro; Tamiya, Eiichi

    2015-12-15

    Polymerase chain reaction (PCR)-based genetic testing has become a routine part of clinical diagnoses and food testing. In these fields, rapid, easy-to-use, and cost-efficient PCR chips are expected to be appeared for providing such testing on-site. In this study, a new autonomous disposable plastic microfluidic PCR chip was created, and was utilized for quantitative detection of pathogenic microorganisms. To control the capillary flow of the following solution in the PCR microchannel, a driving microchannel was newly designed behind the PCR microchannel. This allowed the effective PCR by simply dropping the PCR solution onto the inlet without any external pumps. In order to achieve disposability, injection-molded cyclo-olefin polymer (COP) of a cost-competitive plastic was used for the PCR chip. We discovered that coating the microchannel walls with non-ionic surfactant produced a suitable hydrophilic surface for driving the capillary flow through the 1250-mm long microchannel. As a result, quantitative real-time PCR with the lowest initial concentration of human, Escherichia coli (E. coli), and pathogenic E. coli O157 genomic DNA of 4, 0.0019, 0.031 pg/μl, respectively, was successfully achieved in less than 18 min. Our results indicate that the platform presented in this study provided a rapid, easy-to-use, and low-cost real-time PCR system that could be potentially used for on-site gene testing. PMID:26210470

  1. Imaging Spectrometer on a Chip

    NASA Technical Reports Server (NTRS)

    Wang, Yu; Pain, Bedabrata; Cunningham, Thomas; Zheng, Xinyu

    2007-01-01

    A proposed visible-light imaging spectrometer on a chip would be based on the concept of a heterostructure comprising multiple layers of silicon-based photodetectors interspersed with long-wavelength-pass optical filters. In a typical application, this heterostructure would be replicated in each pixel of an image-detecting integrated circuit of the active-pixel-sensor type (see figure). The design of the heterostructure would exploit the fact that within the visible portion of the spectrum, the characteristic depth of penetration of photons increases with wavelength. Proceeding from the front toward the back, each successive long-wavelength-pass filter would have a longer cutoff wavelength, and each successive photodetector would be made thicker to enable it to absorb a greater proportion of incident longer-wavelength photons. Incident light would pass through the first photodetector and encounter the first filter, which would reflect light having wavelengths shorter than its cutoff wavelength and pass light of longer wavelengths. A large portion of the incident and reflected shorter-wavelength light would be absorbed in the first photodetector. The light that had passed through the first photodetector/filter pair of layers would pass through the second photodetector and encounter the second filter, which would reflect light having wavelengths shorter than its cutoff wavelength while passing light of longer wavelengths. Thus, most of the light reflected by the second filter would lie in the wavelength band between the cutoff wavelengths of the first and second filters. Thus, further, most of the light absorbed in the second photodetector would lie in this wavelength band. In a similar manner, each successive photodetector would detect, predominantly, light in a successively longer wavelength band bounded by the shorter cutoff wavelength of the preceding filter and the longer cutoff wavelength of the following filter.

  2. Investigation of formation mechanisms of chips in orthogonal cutting process

    NASA Astrophysics Data System (ADS)

    Ma, W.

    2012-08-01

    This work investigates the formation mechanisms of chips in orthogonal cutting of mild steel and the transformation conditions between various morphology chips. It is supposed that the modeling material follows the Johnson-Cook constitutive model. In orthogonal cutting process, both the plastic flow and the instability behaviors of chip materials are caused by the plane strain loadings. Therefore, the general instability behaviors of materials in plane strain state are first analyzed with linear perturbation method and a universal instability criterion is established. Based on the analytical results, the formation mechanisms of chips and the transformation conditions between continuous and serrated chips are further studied by instability phase diagram method. The results show that the chip formation strongly depends on the intensity ratios between shear and normal stresses. The ratios of dissipative rates of plastic work done by compression and shear stresses govern the transformation from continuous to serrated chips. These results are verified by the numerical simulations on the orthogonal cutting process.

  3. Modulated Tool-Path (MTP) Chip Breaking System

    SciTech Connect

    Graham, K. B.

    2010-04-01

    The Modulated Tool-Path (MTP) Chip Breaking System produces user-selectable chip lengths and workpiece finishes and is compatible with any material, workpiece shape, and depth of cut. The MTP chip breaking system consistently creates the desired size of chips regardless of workpiece size, shape, or material, and the machine operator does not need to make any adjustments during the machining operation. The system's programmer configures the part program that commands the machine tool to move in a specific fashion to deliver the desired part size, shape, chip length, and workpiece surface finish. The MTP chip breaking system helps manufacturers avoid the detrimental effects of continuous chips, including expensive repair costs, delivery delays, and hazards to personnel.

  4. Mapping Transcription Factors on Extended DNA: A Single Molecule Approach

    NASA Astrophysics Data System (ADS)

    Ebenstein, Yuval; Gassman, Natalie; Weiss, Shimon

    The ability to determine the precise loci and distribution of nucleic acid binding proteins is instrumental to our detailed understanding of cellular processes such as transcription, replication, and chromatin reorganization. Traditional molecular biology approaches and above all Chromatin immunoprecipitation (ChIP) based methods have provided a wealth of information regarding protein-DNA interactions. Nevertheless, existing techniques can only provide average properties of these interactions, since they are based on the accumulation of data from numerous protein-DNA complexes analyzed at the ensemble level. We propose a single molecule approach for direct visualization of DNA binding proteins bound specifically to their recognition sites along a long stretch of DNA such as genomic DNA. Fluorescent Quantum dots are used to tag proteins bound to DNA, and the complex is deposited on a glass substrate by extending the DNA to a linear form. The sample is then imaged optically to determine the precise location of the protein binding site. The method is demonstrated by detecting individual, Quantum dot tagged T7-RNA polymerase enzymes on the bacteriophage T7 genomic DNA and assessing the relative occupancy of the different promoters.

  5. Simulating the Effect of Modulated Tool-Path Chip Breaking On Surface Texture and Chip Length

    SciTech Connect

    Smith, K.S.; McFarland, J.T.; Tursky, D. A.; Assaid, T. S.; Barkman, W. E.; Babelay, Jr., E. F.

    2010-04-30

    One method for creating broken chips in turning processes involves oscillating the cutting tool in the feed direction utilizing the CNC machine axes. The University of North Carolina at Charlotte and the Y-12 National Security Complex have developed and are refining a method to reliably control surface finish and chip length based on a particular machine's dynamic performance. Using computer simulations it is possible to combine the motion of the machine axes with the geometry of the cutting tool to predict the surface characteristics and map the surface texture for a wide range of oscillation parameters. These data allow the selection of oscillation parameters to simultaneously ensure broken chips and acceptable surface characteristics. This paper describes the machine dynamic testing and characterization activities as well as the computational method used for evaluating and predicting chip length and surface texture.

  6. DNA mimicry by proteins.

    PubMed

    Dryden, D T F; Tock, M R

    2006-04-01

    It has been discovered recently, via structural and biophysical analyses, that proteins can mimic DNA structures in order to inhibit proteins that would normally bind to DNA. Mimicry of the phosphate backbone of DNA, the hydrogen-bonding properties of the nucleotide bases and the bending and twisting of the DNA double helix are all present in the mimics discovered to date. These mimics target a range of proteins and enzymes such as DNA restriction enzymes, DNA repair enzymes, DNA gyrase and nucleosomal and nucleoid-associated proteins. The unusual properties of these protein DNA mimics may provide a foundation for the design of targeted inhibitors of DNA-binding proteins. PMID:16545103

  7. GeoChip 4: a functional gene-array-based high-throughput environmental technology for microbial community analysis.

    PubMed

    Tu, Qichao; Yu, Hao; He, Zhili; Deng, Ye; Wu, Liyou; Van Nostrand, Joy D; Zhou, Aifen; Voordeckers, James; Lee, Yong-Jin; Qin, Yujia; Hemme, Christopher L; Shi, Zhou; Xue, Kai; Yuan, Tong; Wang, Aijie; Zhou, Jizhong

    2014-09-01

    Micro-organisms play critical roles in many important biogeochemical processes in the Earth's biosphere. However, understanding and characterizing the functional capacity of microbial communities are still difficult due to the extremely diverse and often uncultivable nature of most micro-organisms. In this study, we developed a new functional gene array, GeoChip 4, for analysing the functional diversity, composition, structure, metabolic potential/activity and dynamics of microbial communities. GeoChip 4 contained approximately 82 000 probes covering 141 995 coding sequences from 410 functional gene families related to microbial carbon (C), nitrogen (N), sulphur (S), and phosphorus (P) cycling, energy metabolism, antibiotic resistance, metal resistance/reduction, organic remediation, stress responses, bacteriophage and virulence. A total of 173 archaeal, 4138 bacterial, 404 eukaryotic and 252 viral strains were targeted, providing the ability to analyse targeted functional gene families of micro-organisms included in all four domains. Experimental assessment using different amounts of DNA suggested that as little as 500 ng environmental DNA was required for good hybridization, and the signal intensities detected were well correlated with the DNA amount used. GeoChip 4 was then applied to study the effect of long-term warming on soil microbial communities at a Central Oklahoma site, with results indicating that microbial communities respond to long-term warming by enriching carbon degradation, nutrient cycling (nitrogen and phosphorous) and stress response gene families. To the best of our knowledge, GeoChip 4 is the most comprehensive functional gene array for microbial community analysis.

  8. Microfluidic Chip-Based Detection and Intraspecies Strain Discrimination of Salmonella Serovars Derived from Whole Blood of Septic Mice

    PubMed Central

    Patterson, Adriana S.; Heithoff, Douglas M.; Ferguson, Brian S.; Soh, H. Tom; Mahan, Michael J.

    2013-01-01

    Salmonella is a zoonotic pathogen that poses a considerable public health and economic burden in the United States and worldwide. Resultant human diseases range from enterocolitis to bacteremia to sepsis and are acutely dependent on the particular serovar of Salmonella enterica subsp. enterica, which comprises over 99% of human-pathogenic S. enterica isolates. Point-of-care methods for detection and strain discrimination of Salmonella serovars would thus have considerable benefit to medical, veterinary, and field applications that safeguard public health and reduce industry-associated losses. Here we describe a single, disposable microfluidic chip that supports isothermal amplification and sequence-specific detection and discrimination of Salmonella serovars derived from whole blood of septic mice. The integrated microfluidic electrochemical DNA (IMED) chip consists of an amplification chamber that supports loop-mediated isothermal amplification (LAMP), a rapid, single-temperature amplification method as an alternative to PCR that offers advantages in terms of sensitivity, reaction speed, and amplicon yield. The amplification chamber is connected via a microchannel to a detection chamber containing a reagentless, multiplexed (here biplex) sensing array for sequence-specific electrochemical DNA (E-DNA) detection of the LAMP products. Validation of the IMED device was assessed by the detection and discrimination of S. enterica subsp. enterica serovars Typhimurium and Choleraesuis, the causative agents of enterocolitis and sepsis in humans, respectively. IMED chips conferred rapid (under 2 h) detection and discrimination of these strains at clinically relevant levels (<1,000 CFU/ml) from whole, unprocessed blood collected from septic animals. The IMED-based chip assay shows considerable promise as a rapid, inexpensive, and portable point-of-care diagnostic platform for the detection and strain-specific discrimination of microbial pathogens. PMID:23354710

  9. A versatile snap chip for high-density sub-nanoliter chip-to-chip reagent transfer

    PubMed Central

    Li, Huiyan; Munzar, Jeffrey D.; Ng, Andy; Juncker, David

    2015-01-01

    The coordinated delivery of minute amounts of different reagents is important for microfluidics and microarrays, but is dependent on advanced equipment such as microarrayers. Previously, we developed the snap chip for the direct transfer of reagents, thus realizing fluidic operations by only manipulating microscope slides. However, owing to the misalignment between arrays spotted on different slides, millimeter spacing was needed between spots and the array density was limited. In this work, we have developed a novel double transfer method and have transferred 625 spots cm−2, corresponding to >10000 spots for a standard microscope slide. A user-friendly snapping system was manufactured to make liquid handling straightforward. Misalignment, which for direct transfer ranged from 150–250 μm, was reduced to <40 μm for double transfer. The snap chip was used to quantify 50 proteins in 16 samples simultaneously, yielding limits of detection in the pg/mL range for 35 proteins. The versatility of the snap chip is illustrated with a 4-plex homogenous enzyme inhibition assay analyzing 128 conditions with precise timing. The versatility and high density of the snap chip with double transfer allows for the development of high throughput reagent transfer protocols compatible with a variety of applications. PMID:26148566

  10. DNA Extraction by Isotachophoresis in a Microfluidic Channel

    SciTech Connect

    Stephenson, S J

    2011-08-10

    Biological assays have many applications. For example, forensics personnel and medical professionals use these tests to diagnose diseases and track their progression or identify pathogens and the host response to them. One limitation of these tests, however, is that most of them target only one piece of the sample - such as bacterial DNA - and other components (e.g. host genomic DNA) get in the way, even though they may be useful for different tests. To address this problem, it would be useful to extract several different substances from a complex biological sample - such as blood - in an inexpensive and efficient manner. This summer, I worked with Maxim Shusteff at Lawrence Livermore National Lab on the Rapid Automated Sample Prep project. The goal of the project is to solve the aforementioned problem by creating a system that uses a series of different extraction methods to extract cells, bacteria, and DNA from a complex biological sample. Biological assays can then be run on purified output samples. In this device, an operator could input a complex sample such as blood or saliva, and would receive separate outputs of cells, bacteria, viruses, and DNA. I had the opportunity to work this summer with isotachophoresis (ITP), a technique that can be used to extract nucleic acids from a sample. This technique is intended to be the last stage of the purification device. Isotachophoresis separates particles based on different electrophoretic mobilities. This technique is convenient for out application because free solution DNA mobility is approximately equal for DNA longer than 300 base pairs in length. The sample of interest - in our case DNA - is fed into the chip with streams of leading electrolyte (LE) and trailing electrolyte (TE). When an electric field is applied, the species migrate based on their electrophoretic mobilities. Because the ions in the leading electrolyte have a high electrophoretic mobility, they race ahead of the slower sample and trailing

  11. DNA Microarray Platform for Detection and Surveillance of Viruses Transmitted by Small Mammals and Arthropods

    PubMed Central

    Khan, Mohd Jaseem; Trabuco, Amanda Cristina; Alfonso, Helda Liz; Figueiredo, Mario Luis; Batista, Weber Cheli; Badra, Soraya Jabur; Figueiredo, Luiz Tadeu; Lavrador, Marco Aurélio

    2016-01-01

    Viruses transmitted by small mammals and arthropods serve as global threats to humans. Most emergent and re-emergent viral agents are transmitted by these groups; therefore, the development of high-throughput screening methods for the detection and surveillance of such viruses is of great interest. In this study, we describe a DNA microarray platform that can be used for screening all viruses transmitted by small mammals and arthropods (SMAvirusChip) with nucleotide sequences that have been deposited in the GenBank. SMAvirusChip was designed with more than 15,000 oligonucleotide probes (60-mers), including viral and control probes. Two SMAvirusChip versions were designed: SMAvirusChip v1 contains 4209 viral probes for the detection of 409 viruses, while SMAvirusChip v2 contains 4943 probes for the detection of 416 viruses. SMAvirusChip was evaluated with 20 laboratory reference-strain viruses. These viruses could be specifically detected when alone in a sample or when artificially mixed within a single sample. The sensitivity of SMAvirusChip was evaluated using 10-fold serial dilutions of dengue virus (DENV). The results showed a detection limit as low as 2.6E3 RNA copies/mL. Additionally, the sensitivity was one log10 lower (2.6E2 RNA copies/mL) than quantitative real-time RT-PCR and sufficient to detect viral genomes in clinical samples. The detection of DENV in serum samples of DENV-infected patients (n = 6) and in a whole blood sample spiked with DENV confirmed the applicability of SMAvirusChip for the detection of viruses in clinical samples. In addition, in a pool of mosquito samples spiked with DENV, the virus was also detectable. SMAvirusChip was able to specifically detect viruses in cell cultures, serum samples, total blood samples and a pool of mosquitoes, confirming that cellular RNA/DNA did not interfere with the assay. Therefore, SMAvirusChip may represent an innovative surveillance method for the rapid identification of viruses transmitted by small

  12. Chromatin Immunoprecipitation Assay for the Identification of Arabidopsis Protein-DNA Interactions In Vivo.

    PubMed

    Komar, Dorota N; Mouriz, Alfonso; Jarillo, José A; Piñeiro, Manuel

    2016-01-14

    Intricate gene regulatory networks orchestrate biological processes and developmental transitions in plants. Selective transcriptional activation and silencing of genes mediate the response of plants to environmental signals and developmental cues. Therefore, insights into the mechanisms that control plant gene expression are essential to gain a deep understanding of how biological processes are regulated in plants. The chromatin immunoprecipitation (ChIP) technique described here is a procedure to identify the DNA-binding sites of proteins in genes or genomic regions of the model species Arabidopsis thaliana. The interactions with DNA of proteins of interest such as transcription factors, chromatin proteins or posttranslationally modified versions of histones can be efficiently analyzed with the ChIP protocol. This method is based on the fixation of protein-DNA interactions in vivo, random fragmentation of chromatin, immunoprecipitation of protein-DNA complexes with specific antibodies, and quantification of the DNA associated with the protein of interest by PCR techniques. The use of this methodology in Arabidopsis has contributed significantly to unveil transcriptional regulatory mechanisms that control a variety of plant biological processes. This approach allowed the identification of the binding sites of the Arabidopsis chromatin protein EBS to regulatory regions of the master gene of flowering FT. The impact of this protein in the accumulation of particular histone marks in the genomic region of FT was also revealed through ChIP analysis.

  13. Stem-end chip defect: trial summary from 2009-2012

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Stem-end chip defect is characterized by dark fried color along the vascular and adjacent tissues at the basal (stem) end of potato chips. Stem-end defect chips are unacceptable to chip processors and stem-end defect tubers may be rejected at processing plants. Stem-end chip defect occurs erraticall...

  14. DNA ligase I, the replicative DNA ligase

    PubMed Central

    Howes, Timothy R.L.; Tomkinson, Alan E.

    2013-01-01

    Multiple DNA ligation events are required to join the Okazaki fragments generated during lagging strand DNA synthesis. In eukaryotes, this is primarily carried out by members of the DNA ligase I family. The C-terminal catalytic region of these enzymes is composed of three domains: a DNA binding domain, an adenylation domain and an OB-fold domain. In the absence of DNA, these domains adopt an extended structure but transition into a compact ring structure when they engage a DNA nick, with each of the domains contacting the DNA. The non-catalytic N-terminal region of eukaryotic DNA ligase I is responsible for the specific participation of these enzymes in DNA replication. This proline-rich unstructured region contains the nuclear localization signal and a PCNA interaction motif that is critical for localization to replication foci and efficient joining of Okazaki fragments. DNA ligase I initially engages the PCNA trimer via this interaction motif which is located at the extreme N-terminus of this flexible region. It is likely that this facilitates an additional interaction between the DNA binding domain and the PCNA ring. The similar size and shape of the rings formed by the PCNA trimer and the DNA ligase I catalytic region when it engages a DNA nick suggest that these proteins interact to form a double-ring structure during the joining of Okazaki fragments. DNA ligase I also interacts with replication factor C, the factor that loads the PCNA trimeric ring onto DNA. This interaction, which is regulated by phosphorylation of the non-catalytic N-terminus of DNA ligase I, also appears to be critical for DNA replication. PMID:22918593

  15. From microfluidic modules to an integrated Lab-on-a-chip system for the detection of Francisella tularensis

    NASA Astrophysics Data System (ADS)

    Hlawatsch, Nadine; Krumbholz, Marco; Prüfer, Anna; Moche, Christian; Becker, Holger; Gärtner, Claudia

    2013-05-01

    Lab-on-a-chip (LoC) systems translating the whole process of pathogen analysis to an integrated, miniaturized, and automatically functioning microfluidic platform are generally expected to be very promising future diagnostic approaches. The development of such a LoC system for the detection of bacterial pathogens applied to the example pathogen Francisella tularensis is described in this report. To allow functional testing of the whole process cascade before final device integration, various bio-analytical steps such as cell lysis, DNA extraction and purification, continuous-flow PCR and analyte detection have been adapted to unique functional microfluidic modules. As a successive step, positively tested modules for pathogen detection have been successfully assembled to an integrated chip. Moreover, technical solutions for a smooth interaction between sample input from the outer world as well as microfluidic chip and chip driving instrument have been developed. In conclusion, a full repertoire of analytical tools have been developed and successfully tested in the concerted manner of a functionally integrated microfluidic device representing a tool for future diagnostic approaches.

  16. Towards a cellular multi-parameter analysis platform: fluorescence in situ hybridization (FISH) on microhole-array chips.

    PubMed

    Kurz, Christian M; Moosdijk, Stefan V D; Thielecke, Hagen; Velten, Thomas

    2011-01-01

    Highly-sensitive analysis systems based on cellular multi-parameter are needed in the diagnostics. Therefore we improved our previously developed chip platform for another additional analysis method, the fluorescence in situ hybridization. Fluorescence in situ hybridization (FISH) is a technique used in the diagnostics to determine the localization and the presence or absence of specific DNA sequence. To improve this labor- and cost-intensive method, we reduced the assay consumption by a factor of 5 compared to the standard protocol. Microhole chips were used for making the cells well addressable. The chips were fabricated by semiconductor technology on the basis of a Silicon wafer with a thin deposited silicon nitride layer (Si(3)N(4)). Human retina pigment epithelia (ARPE-19) cells were arrayed on 5-μm holes of a 35 × 35 microhole-array by a gently negative differential pressure of around 5 mbar. After 3 hours of incubation the cells were attached to the chip and the FISH protocol was applied to the positioned cells. A LabView software was developed to simplify the analysis. The software automatically counts the number of dots (positive labeled chromosome regions) as well as the distance between adjacent dots. Our developed platform reduces the assay consumption and the labor time. Furthermore, during the 3 hours of incubation non-invasive or minimal-invasive methods like Raman- and impedance-spectroscopy can be applied. PMID:22256298

  17. Genome-wide analysis of DNA methylation in hepatoblastoma tissues

    PubMed Central

    Cui, Ximao; Liu, Baihui; Zheng, Shan; Dong, Kuiran; Dong, Rui

    2016-01-01

    DNA methylation has a crucial role in cancer biology. In the present study, a genome-wide analysis of DNA methylation in hepatoblastoma (HB) tissues was performed to verify differential methylation levels between HB and normal tissues. As alpha-fetoprotein (AFP) has a critical role in HB, AFP methylation levels were also detected using pyrosequencing. Normal and HB liver tissue samples (frozen tissue) were obtained from patients with HB. Genome-wide analysis of DNA methylation in these tissues was performed using an Infinium HumanMethylation450 BeadChip, and the results were confirmed with reverse transcription-quantitative polymerase chain reaction. The Infinium HumanMethylation450 BeadChip demonstrated distinctively less methylation in HB tissues than in non-tumor tissues. In addition, methylation enrichment was observed in positions near the transcription start site of AFP, which exhibited lower methylation levels in HB tissues than in non-tumor liver tissues. Lastly, a significant negative correlation was observed between AFP messenger RNA expression and DNA methylation percentage, using linear Pearson's R correlation coefficients. The present results demonstrate differential methylation levels between HB and normal tissues, and imply that aberrant methylation of AFP in HB could reflect HB development. Expansion of these findings could provide useful insight into HB biology. PMID:27446465

  18. [Species identification of grouper and snapper in Taiwan Strait using polymerase chain reaction-restriction fragment length polymorphism analysis and lab-on-a-chip system].

    PubMed

    Chen, Shuangya; Zhang, Jin; Chen, Weiling; Xu, Dunming; Zhou, Yu

    2011-07-01

    Polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) analysis and lab-on-a-chip system were used to identify grouper and snapper species in Taiwan Strait. A fragment of 464 bp length of mitochondrial cytochrome b gene was amplified by PCR and the products were digested with restriction enzymes Dde I , Hae III and NLa III, individually. The fragments generated after digestion were further resolved on the DNA Chip. Eight grouper species and five snapper species were successfully identified. The results demonstrated that PCR-RFLP analysis and lab-on-a-chip system provide a fast, easy, automated, and reliable analysis approach. This approach is potential for the purpose of fish adulteration control.

  19. Dopamine-functionalized InP/ZnS quantum dots as fluorescence probes for the detection of adenosine in microfluidic chip

    PubMed Central

    Ankireddy, Seshadri Reddy; Kim, Jongsung

    2015-01-01

    Microbeads are frequently used as solid supports for biomolecules such as proteins and nucleic acids in heterogeneous microfluidic assays. Chip-based, quantum dot (QD)-bead-biomolecule probes have been used for the detection of various types of DNA. In this study, we developed dopamine (DA)-functionalized InP/ZnS QDs (QDs-DA) as fluorescence probes for the detection of adenosine in microfluidic chips. The photoluminescence (PL) intensity of the QDs-DA is quenched by Zn2+ because of the strong coordination interactions. In the presence of adenosine, Zn2+ cations preferentially bind to adenosine, and the PL intensity of the QDs-DA is recovered. A polydimethylsiloxane-based microfluidic chip was fabricated, and adenosine detection was confirmed using QDs-DA probes. PMID:26347351

  20. Single-nucleotide polymorphism genotyping on optical thin-film biosensor chips

    PubMed Central

    Zhong, Xiao-bo; Reynolds, Robert; Kidd, Judith R.; Kidd, Kenneth K.; Jenison, Robert; Marlar, Richard A.; Ward, David C.

    2003-01-01

    Single-nucleotide polymorphisms (SNPs) constitute the bulk of human genetic variation and provide excellent markers to identify genetic factors contributing to complex disease susceptibility. A rapid, sensitive, and inexpensive assay is important for large-scale SNP scoring. Here we report the development of a multiplex SNP detection system using silicon chips coated to create a thin-film optical biosensor. Allele-discriminating, aldehyde-labeled oligonucleotides are arrayed and covalently attached to a hydrazinederivatized chip surface. Target sequences (e.g., PCR amplicons) then are hybridized in the presence of a mixture of biotinylated detector probes, one for each SNP, and a thermostable DNA ligase. After a stringent wash (0.01 M NaOH), ligation of biotinylated detector probes to perfectly matched capture oligomers is visualized as a color change on the chip surface (gold to blue/purple) after brief incubations with an anti-biotin IgG-horseradish peroxidase conjugate and a precipitable horseradish peroxidase substrate. Testing of PCR fragments is completed in 30–40 min. Up to several hundred SNPs can be assayed on a 36-mm2 chip, and SNP scoring can be done by eye or with a simple digital-camera system. This assay is extremely robust, exhibits high sensitivity and specificity, and is format-flexible and economical. In studies of mutations associated with risk for venous thrombosis and genotyping/haplotyping of African-American samples, we document high-fidelity analysis with 0 misassignments in 500 assays performed in duplicate. PMID:12975525

  1. Infrared temperature control system for a completely noncontact polymerase chain reaction in microfluidic chips.

    PubMed

    Roper, Michael G; Easley, Christopher J; Legendre, Lindsay A; Humphrey, Joseph A C; Landers, James P

    2007-02-15

    A completely noncontact temperature system is described for amplification of DNA via the polymerase chain reaction (PCR) in glass microfluidic chips. An infrared (IR)-sensitive pyrometer was calibrated against a thermocouple inserted into a 550-nL PCR chamber and used to monitor the temperature of the glass surface above the PCR chamber during heating and cooling induced by a tungsten lamp and convective air source, respectively. A time lag of less than 1 s was observed between maximum heating rates of the solution and surface, indicating that thermal equilibrium was attained rapidly. Moreover, the time lag was corroborated using a one-dimensional heat-transfer model, which provided insight into the characteristics of the device and environment that caused the time lag. This knowledge will, in turn, allow for future tailoring of the devices to specific applications. To alleviate the need for calibrating the pyrometer with a thermocouple, the on-chip calibration of pyrometer was accomplished by sensing the boiling of two solutions, water and an azeotrope, and comparing the pyrometer output voltage against the known boiling points of these solutions. The "boiling point calibration" was successful as indicated by the subsequent chip-based IR-PCR amplification of a 211-bp fragment of the B. anthracis genome in a chamber reduced beyond the dimensions of a thermocouple. To improve the heating rates, a parabolic gold mirror was positioned above the microfluidic chip, which expedited PCR amplification to 18.8 min for a 30-cycle, three-temperature protocol. PMID:17297927

  2. A programmable and reconfigurable microfluidic chip.

    PubMed

    Renaudot, Raphael; Agache, Vincent; Fouillet, Yves; Laffite, Guillaume; Bisceglia, Emilie; Jalabert, Laurent; Kumemura, Momoko; Collard, Dominique; Fujita, Hiroyuki

    2013-12-01

    This article reports an original concept enabling the rapid fabrication of continuous-flow microfluidic chips with a programmable and reconfigurable geometry. The concept is based on a digital microfluidic platform featuring an array of individually addressable electrodes. A selection of electrodes is switched on sequentially to create a de-ionized (DI) water finger specific pattern, while the surrounding medium consists of liquid-phase paraffin. The water displacement is induced by both electrowetting on dielectric and liquid dielectrophoresis phenomena. Once the targeted DI water pattern is obtained, the chip temperature is lowered by turning on an integrated thermoelectric cooler, forming channel structures made of solidified paraffin with edges delimitated by the DI water pattern. As a result, the chip can be used afterwards to conduct in-flow continuous microfluidic experiments. This process is resettable and reversible by heating up the chip to melt the paraffin and reconfigure the microchannel design on demand, offering the advantages of cost, adaptability, and robustness. This paper reports experimental results describing the overall concept, which is illustrated with typical and basic fluidic geometries.

  3. Microelectronic Chips For Radiation-Dose Tests

    NASA Technical Reports Server (NTRS)

    Buehler, Martin G.; Lin, Yu-Sang; Ray, Kevin P.; Sokoloski, Martin M.

    1993-01-01

    Custom-made single-chip complementary metal-oxide semiconductor (CMOS) integrated circuit designed to reveal effects of ionizing radiation on itself and similar integrated circuits. Potential terrestrial use: safety-oriented monitoring of ionizing radiation at nuclear powerplants, nuclear-waste sites, and the like.

  4. System-on-Chip Design and Implementation

    ERIC Educational Resources Information Center

    Brackenbury, L. E. M.; Plana, L. A.; Pepper, J.

    2010-01-01

    The system-on-chip module described here builds on a grounding in digital hardware and system architecture. It is thus appropriate for third-year undergraduate computer science and computer engineering students, for post-graduate students, and as a training opportunity for post-graduate research students. The course incorporates significant…

  5. Hybrid photonic chip interferometer for embedded metrology

    NASA Astrophysics Data System (ADS)

    Kumar, P.; Martin, H.; Maxwell, G.; Jiang, X.

    2014-03-01

    Embedded metrology is the provision of metrology on the manufacturing platform, enabling measurement without the removal of the work piece. Providing closer integration of metrology upon the manufacturing platform can lead to the better control and increased throughput. In this work we present the development of a high precision hybrid optical chip interferometer metrology device. The complete metrology sensor system is structured into two parts; optical chip and optical probe. The hybrid optical chip interferometer is based on a silica-on-silicon etched integrated-optic motherboard containing waveguide structures and evanescent couplers. Upon the motherboard, electro-optic components such as photodiodes and a semiconductor gain block are mounted and bonded to provide the required functionality. The key structure in the device is a tunable laser module based upon an external-cavity diode laser (ECDL). Within the cavity is a multi-layer thin film filter which is rotated to select the longitudinal mode at which the laser operates. An optical probe, which uses a blazed diffracting grating and collimating objective lens, focuses light of different wavelengths laterally over the measurand. Incident laser light is then tuned in wavelength time to effectively sweep an `optical stylus' over the surface. Wavelength scanning and rapid phase shifting can then retrieve the path length change and thus the surface height. We give an overview of the overall design of the final hybrid photonic chip interferometer, constituent components, device integration and packaging as well as experimental test results from the current version now under evaluation.

  6. Programmable Electro Osmotic Lab on a Chip

    NASA Astrophysics Data System (ADS)

    Class, Andreas G.

    2014-11-01

    We propose to use a 2D check-board patterned surface with alternating zeta potential made of semiconductors and individually controllable electrodes surrounding each field to drive by electro osmosis an arbitrary flow along the surface within the cavity of a lab-on-a-chip. In contrast to other fluid mechanic devices the flow is not driven by pressure gradients but rather by a controllable fluid velocity within the Debay boundary layer. Thus fluid is transported like a parcel on a conveyor belt. The use of alternating zeta potential fields and alternating electrode polarities allows to transport flow along multiple fields without the need to increase voltage. Basic functionality of the chip is accomplished by appropriate programming: fluid transport along straight and curved path, merging and splitting flow paths, flow crossing by red light traffic control, and mixing. Implementing sensors for electric resistance on the Lab-On-A-Chip allows to program a diagnosis application using electrophoresis for detection. Transport within the Lab-On-A-Chip can be described by Stokes-flow subject to the boundary conditions given by asymptotic theory in the thin-Debay-layer-limit describing field driven electro kinetic effects.

  7. Writing for a Change, Writing for Chip

    ERIC Educational Resources Information Center

    Berry, Patrick W.

    2014-01-01

    What does it mean to write for change? How do we negotiate the space between hope and critique? Drawing on Dewey's notion of a common faith, this article contemplates what the author learned from Chip Bruce. It suggests that when we compartmentalize the ideal and the everyday, the hopeful and the critical, we reduce the complexity of human…

  8. Flip-a-Chip to Build Vocabulary.

    ERIC Educational Resources Information Center

    Mountain, Lee

    2002-01-01

    Presents a word-game strategy that builds vocabulary and comprehension while motivating students. Concludes that activities like Flip-a-Chip (along with crossword puzzles and other forms of wordplay) have helped the author create a pleasantly literate environment in her classroom. (SG)

  9. Light-colored, Low Acrylamide Potato Chips

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Potato tubers are stored at cold temperatures to prevent sprouting, minimize disease losses and increase the marketing window. Cold storage also causes an accumulation of reducing sugars, a phenomenon referred to as cold-induced sweetening. Unacceptable, dark colored chips and fries are formed durin...

  10. The V-Chip--Victory or Vendetta?

    ERIC Educational Resources Information Center

    Payne, Ron

    1997-01-01

    Parents can install the v-chip microchip in their televisions to block out programs high in violence, sex, or other objectional material. Examines the views of supporters, who see it as a coping tool for the information age and of detractors who see it as an affront to the First Amendment guarantee of free speech. (SM)

  11. Increasing security in inter-chip communication

    DOEpatents

    Edwards, Nathan J; Hamlet, Jason; Bauer, Todd; Helinski, Ryan

    2014-10-28

    An apparatus for increasing security in inter-chip communication includes a sending control module, a communication bus, and a receiving control module. The communication bus is coupled between the sending control module and the receiving control module. The sending control module operates to send data on the communication bus, disable the communication bus when threats are detected, or both.

  12. Sensing systems using chip-based spectrometers

    NASA Astrophysics Data System (ADS)

    Nitkowski, Arthur; Preston, Kyle J.; Sherwood-Droz, Nicolás.; Behr, Bradford B.; Bismilla, Yusuf; Cenko, Andrew T.; DesRoches, Brandon; Meade, Jeffrey T.; Munro, Elizabeth A.; Slaa, Jared; Schmidt, Bradley S.; Hajian, Arsen R.

    2014-06-01

    Tornado Spectral Systems has developed a new chip-based spectrometer called OCTANE, the Optical Coherence Tomography Advanced Nanophotonic Engine, built using a planar lightwave circuit with integrated waveguides fabricated on a silicon wafer. While designed for spectral domain optical coherence tomography (SD-OCT) systems, the same miniaturized technology can be applied to many other spectroscopic applications. The field of integrated optics enables the design of complex optical systems which are monolithically integrated on silicon chips. The form factors of these systems can be significantly smaller, more robust and less expensive than their equivalent free-space counterparts. Fabrication techniques and material systems developed for microelectronics have previously been adapted for integrated optics in the telecom industry, where millions of chip-based components are used to power the optical backbone of the internet. We have further adapted the photonic technology platform for spectroscopy applications, allowing unheard-of economies of scale for these types of optical devices. Instead of changing lenses and aligning systems, these devices are accurately designed programmatically and are easily customized for specific applications. Spectrometers using integrated optics have large advantages in systems where size, robustness and cost matter: field-deployable devices, UAVs, UUVs, satellites, handheld scanning and more. We will discuss the performance characteristics of our chip-based spectrometers and the type of spectral sensing applications enabled by this technology.

  13. DNA modifications: Another stable base in DNA

    NASA Astrophysics Data System (ADS)

    Brazauskas, Pijus; Kriaucionis, Skirmantas

    2014-12-01

    Oxidation of 5-methylcytosine has been proposed to mediate active and passive DNA demethylation. Tracking the history of DNA modifications has now provided the first solid evidence that 5-hydroxymethylcytosine is a stable epigenetic modification.

  14. Micromachined polymerase chain reaction system for multiple DNA amplification of upper respiratory tract infectious diseases.

    PubMed

    Liao, Chia-Sheng; Lee, Gwo-Bin; Wu, Jiunn-Jong; Chang, Chih-Ching; Hsieh, Tsung-Min; Huang, Fu-Chun; Luo, Ching-Hsing

    2005-01-15

    This paper presents a micro polymerase chain reaction (PCR) chip for the DNA-based diagnosis of microorganism genes and the detection of their corresponding antibiotic-resistant genes. The micro PCR chip comprises cheap biocompatible soda-lime glass substrates with integrated thin-film platinum resistors as heating/sensing elements, and is fabricated using micro-electro-mechanical-system (MEMS) techniques in a reliable batch-fabrication process. The heating and temperature sensing elements are made of the same material and are located inside the reaction chamber in order to ensure a uniform temperature distribution. This study performs the detection of several genes associated with upper respiratory tract infection microorganisms, i.e. Streptococcus pneumoniae, Haemopilus influenze, Staphylococcu aureus, Streptococcus pyogenes, and Neisseria meningitides, together with their corresponding antibiotic-resistant genes. The lower thermal inertia of the proposed micro PCR chip relative to conventional bench-top PCR systems enables a more rapid detection operation with reduced sample and reagent consumption. The experimental data reveal that the high heating and cooling rates of the system (20 and 10 degrees C/s, respectively) permit successful DNA amplification within 15 min. The micro PCR chip is also capable of performing multiple DNA amplification, i.e. the simultaneous duplication of multiple genes under different conditions in separate reaction wells. Compared with the large-scale PCR system, it is greatly advantageous for fast diagnosis of multiple infectious diseases. Multiplex PCR amplification of two DNA segments in the same well is also feasible using the proposed micro device. The developed micro PCR chip provides a crucial tool for genetic analysis, molecular biology, infectious disease detection, and many other biomedical applications. PMID:15590288

  15. Sperm DNA oxidative damage and DNA adducts.

    PubMed

    Jeng, Hueiwang Anna; Pan, Chih-Hong; Chao, Mu-Rong; Lin, Wen-Yi

    2015-12-01

    The objective of this study was to investigate DNA damage and adducts in sperm from coke oven workers who have been exposed to polycyclic aromatic hydrocarbons. A longitudinal study was conducted with repeated measurements during spermatogenesis. Coke-oven workers (n=112) from a coke-oven plant served the PAH-exposed group, while administrators and security personnel (n=67) served the control. Routine semen parameters (concentration, motility, vitality, and morphology) were analyzed simultaneously; the assessment of sperm DNA integrity endpoints included DNA fragmentation, bulky DNA adducts, and 8-oxo-7,8-dihydro-2'-deoxyguanosine (8-oxo-dGuo). The degree of sperm DNA fragmentation was measured using the terminal deoxynucleotidyl transferase-mediated dUTP nick end-labeling (TUNEL) assay and sperm chromatin structure assay (SCSA). The PAH-exposed group had a significant increase in bulky DNA adducts and 8-oxo-dGuo compared to the control subjects (Ps=0.002 and 0.045, respectively). Coke oven workers' percentages of DNA fragmentation and denaturation from the PAH-exposed group were not significantly different from those of the control subjects (Ps=0.232 and 0.245, respectively). Routine semen parameters and DNA integrity endpoints were not correlated. Concentrations of 8-oxo-dGuo were positively correlated with percentages of DNA fragmentation measured by both TUNEL and SCSA (Ps=0.045 and 0.034, respectively). However, the concentrations of 8-oxo-dGuo and percentages of DNA fragmentation did not correlate with concentrations of bulky DNA adducts. In summary, coke oven workers with chronic exposure to PAHs experienced decreased sperm DNA integrity. Oxidative stress could contribute to the degree of DNA fragmentation. Bulky DNA adducts may be independent of the formation of DNA fragmentation and oxidative adducts in sperm. Monitoring sperm DNA integrity is recommended as a part of the process of assessing the impact of occupational and environmental toxins on sperm.

  16. Sperm DNA oxidative damage and DNA adducts.

    PubMed

    Jeng, Hueiwang Anna; Pan, Chih-Hong; Chao, Mu-Rong; Lin, Wen-Yi

    2015-12-01

    The objective of this study was to investigate DNA damage and adducts in sperm from coke oven workers who have been exposed to polycyclic aromatic hydrocarbons. A longitudinal study was conducted with repeated measurements during spermatogenesis. Coke-oven workers (n=112) from a coke-oven plant served the PAH-exposed group, while administrators and security personnel (n=67) served the control. Routine semen parameters (concentration, motility, vitality, and morphology) were analyzed simultaneously; the assessment of sperm DNA integrity endpoints included DNA fragmentation, bulky DNA adducts, and 8-oxo-7,8-dihydro-2'-deoxyguanosine (8-oxo-dGuo). The degree of sperm DNA fragmentation was measured using the terminal deoxynucleotidyl transferase-mediated dUTP nick end-labeling (TUNEL) assay and sperm chromatin structure assay (SCSA). The PAH-exposed group had a significant increase in bulky DNA adducts and 8-oxo-dGuo compared to the control subjects (Ps=0.002 and 0.045, respectively). Coke oven workers' percentages of DNA fragmentation and denaturation from the PAH-exposed group were not significantly different from those of the control subjects (Ps=0.232 and 0.245, respectively). Routine semen parameters and DNA integrity endpoints were not correlated. Concentrations of 8-oxo-dGuo were positively correlated with percentages of DNA fragmentation measured by both TUNEL and SCSA (Ps=0.045 and 0.034, respectively). However, the concentrations of 8-oxo-dGuo and percentages of DNA fragmentation did not correlate with concentrations of bulky DNA adducts. In summary, coke oven workers with chronic exposure to PAHs experienced decreased sperm DNA integrity. Oxidative stress could contribute to the degree of DNA fragmentation. Bulky DNA adducts may be independent of the formation of DNA fragmentation and oxidative adducts in sperm. Monitoring sperm DNA integrity is recommended as a part of the process of assessing the impact of occupational and environmental toxins on sperm

  17. Synthesis of DNA

    DOEpatents

    Mariella, Jr., Raymond P.

    2008-11-18

    A method of synthesizing a desired double-stranded DNA of a predetermined length and of a predetermined sequence. Preselected sequence segments that will complete the desired double-stranded DNA are determined. Preselected segment sequences of DNA that will be used to complete the desired double-stranded DNA are provided. The preselected segment sequences of DNA are assembled to produce the desired double-stranded DNA.

  18. Electrical detection of single-base DNA mutation using functionalized nanoparticles

    NASA Astrophysics Data System (ADS)

    Noor, Mohammud R.; Goyal, Swati; Christensen, Shawn M.; Iqbal, Samir M.

    2009-08-01

    We report an electrical scheme to detect specific DNA. Engineered hairpin probe DNA are immobilized on a silicon chip between gold nanoelectrodes. Hybridization of target DNA to the hairpin melts the stem nucleotides. Gold nanoparticle-conjugated universal reporter sequence detects the open hairpins by annealing to the exposed stem nucleotides. The gold nanoparticles increase charge conduction between the electrodes. Specifically, we report on a hairpin probe designed to detect a medically relevant mutant form of the K-ras oncogene. Direct current measurements show three orders of magnitude increase in conductivity for as low as 2fmol of target molecules.

  19. Electrical detection of single-base DNA mutation using functionalized nanoparticles

    NASA Astrophysics Data System (ADS)

    Noor, Mohammud R.; Goyal, Swati; Christensen, Shawn M.; Iqbal, Samir M.

    2009-08-01

    We report an electrical scheme to detect specific DNA. Engineered hairpin probe DNA are immobilized on a silicon chip between gold nanoelectrodes. Hybridization of target DNA to the hairpin melts the stem nucleotides. Gold nanoparticle-conjugated universal reporter sequence detects the open hairpins by annealing to the exposed stem nucleotides. The gold nanoparticles increase charge conduction between the electrodes. Specifically, we report on a hairpin probe designed to detect a medically relevant mutant form of the K-ras oncogene. Direct current measurements show three orders of magnitude increase in conductivity for as low as 2 fmol of target molecules.

  20. Genome-wide Mapping Reveals Conservation of Promoter DNA Methylation Following Chicken Domestication

    PubMed Central

    Li, Qinghe; Wang, Yuanyuan; Hu, Xiaoxiang; Zhao, Yaofeng; Li, Ning

    2015-01-01

    It is well-known that environment influences DNA methylation, however, the extent of heritable DNA methylation variation following animal domestication remains largely unknown. Using meDIP-chip we mapped the promoter methylomes for 23,316 genes in muscle tissues of ancestral and domestic chickens. We systematically examined the variation of promoter DNA methylation in terms of different breeds, differentially expressed genes, SNPs and genes undergo genetic selection sweeps. While considerable changes in DNA sequence and gene expression programs were prevalent, we found that the inter-strain DNA methylation patterns were highly conserved in promoter region between the wild and domestic chicken breeds. Our data suggests a global preservation of DNA methylation between the wild and domestic chicken breeds in either a genome-wide or locus-specific scale in chick muscle tissues. PMID:25735894

  1. Genome-wide mapping reveals conservation of promoter DNA methylation following chicken domestication.

    PubMed

    Li, Qinghe; Wang, Yuanyuan; Hu, Xiaoxiang; Zhao, Yaofeng; Li, Ning

    2015-01-01

    It is well-known that environment influences DNA methylation, however, the extent of heritable DNA methylation variation following animal domestication remains largely unknown. Using meDIP-chip we mapped the promoter methylomes for 23,316 genes in muscle tissues of ancestral and domestic chickens. We systematically examined the variation of promoter DNA methylation in terms of different breeds, differentially expressed genes, SNPs and genes undergo genetic selection sweeps. While considerable changes in DNA sequence and gene expression programs were prevalent, we found that the inter-strain DNA methylation patterns were highly conserved in promoter region between the wild and domestic chicken breeds. Our data suggests a global preservation of DNA methylation between the wild and domestic chicken breeds in either a genome-wide or locus-specific scale in chick muscle tissues.

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

    PubMed

    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.

  3. Scalable amplification of strand subsets from chip-synthesized oligonucleotide libraries.

    PubMed

    Schmidt, Thorsten L; Beliveau, Brian J; Uca, Yavuz O; Theilmann, Mark; Da Cruz, Felipe; Wu, Chao-Ting; Shih, William M

    2015-11-16

    Synthetic oligonucleotides are the main cost factor for studies in DNA nanotechnology, genetics and synthetic biology, which all require thousands of these at high quality. Inexpensive chip-synthesized oligonucleotide libraries can contain hundreds of thousands of distinct sequences, however only at sub-femtomole quantities per strand. Here we present a selective oligonucleotide amplification method, based on three rounds of rolling-circle amplification, that produces nanomole amounts of single-stranded oligonucleotides per millilitre reaction. In a multistep one-pot procedure, subsets of hundreds or thousands of single-stranded DNAs with different lengths can selectively be amplified and purified together. These oligonucleotides are used to fold several DNA nanostructures and as primary fluorescence in situ hybridization probes. The amplification cost is lower than other reported methods (typically around US$ 20 per nanomole total oligonucleotides produced) and is dominated by the use of commercial enzymes.

  4. Scalable amplification of strand subsets from chip-synthesized oligonucleotide libraries

    PubMed Central

    Schmidt, Thorsten L.; Beliveau, Brian J.; Uca, Yavuz O.; Theilmann, Mark; Da Cruz, Felipe; Wu, Chao-Ting; Shih, William M.

    2015-01-01

    Synthetic oligonucleotides are the main cost factor for studies in DNA nanotechnology, genetics and synthetic biology, which all require thousands of these at high quality. Inexpensive chip-synthesized oligonucleotide libraries can contain hundreds of thousands of distinct sequences, however only at sub-femtomole quantities per strand. Here we present a selective oligonucleotide amplification method, based on three rounds of rolling-circle amplification, that produces nanomole amounts of single-stranded oligonucleotides per millilitre reaction. In a multistep one-pot procedure, subsets of hundreds or thousands of single-stranded DNAs with different lengths can selectively be amplified and purified together. These oligonucleotides are used to fold several DNA nanostructures and as primary fluorescence in situ hybridization probes. The amplification cost is lower than other reported methods (typically around US$ 20 per nanomole total oligonucleotides produced) and is dominated by the use of commercial enzymes. PMID:26567534

  5. Scalable amplification of strand subsets from chip-synthesized oligonucleotide libraries

    NASA Astrophysics Data System (ADS)

    Schmidt, Thorsten L.; Beliveau, Brian J.; Uca, Yavuz O.; Theilmann, Mark; da Cruz, Felipe; Wu, Chao-Ting; Shih, William M.

    2015-11-01

    Synthetic oligonucleotides are the main cost factor for studies in DNA nanotechnology, genetics and synthetic biology, which all require thousands of these at high quality. Inexpensive chip-synthesized oligonucleotide libraries can contain hundreds of thousands of distinct sequences, however only at sub-femtomole quantities per strand. Here we present a selective oligonucleotide amplification method, based on three rounds of rolling-circle amplification, that produces nanomole amounts of single-stranded oligonucleotides per millilitre reaction. In a multistep one-pot procedure, subsets of hundreds or thousands of single-stranded DNAs with different lengths can selectively be amplified and purified together. These oligonucleotides are used to fold several DNA nanostructures and as primary fluorescence in situ hybridization probes. The amplification cost is lower than other reported methods (typically around US$ 20 per nanomole total oligonucleotides produced) and is dominated by the use of commercial enzymes.

  6. Quantitative Visualization of ChIP-chip Data by Using Linked Views

    SciTech Connect

    Huang, Min-Yu; Weber, Gunther; Li, Xiao-Yong; Biggin, Mark; Hamann, Bernd

    2010-11-05

    Most analyses of ChIP-chip in vivo DNA binding have focused on qualitative descriptions of whether genomic regions are bound or not. There is increasing evidence, however, that factors bind in a highly overlapping manner to the same genomic regions and that it is quantitative differences in occupancy on these commonly bound regions that are the critical determinants of the different biological specificity of factors. As a result, it is critical to have a tool to facilitate the quantitative visualization of differences between transcription factors and the genomic regions they bind to understand each factor's unique roles in the network. We have developed a framework which combines several visualizations via brushing-and-linking to allow the user to interactively analyze and explore in vivo DNA binding data of multiple transcription factors. We describe these visualization types and also provide a discussion of biological examples in this paper.

  7. DNA encoding a DNA repair protein

    DOEpatents

    Petrini, John H.; Morgan, William Francis; Maser, Richard Scott; Carney, James Patrick

    2006-08-15

    An isolated and purified DNA molecule encoding a DNA repair protein, p95, is provided, as is isolated and purified p95. Also provided are methods of detecting p95 and DNA encoding p95. The invention further provides p95 knock-out mice.

  8. High-speed DSP applied to a multimedia chip set

    NASA Astrophysics Data System (ADS)

    Chester, David B.

    1996-06-01

    The hardware portion of the Harris Semiconductor Personal Computer Multimedia System is a 5 chip set which implements, in conjunction with a host processor and associated software and firmware, a complete H.320 video teleconferencing capability over ISDN 2B lines. The chip is comprised of a PAL/NTSC Video Encoder, a PAL/NTSC Video Decoder, a Video Codec, a Bus Interface and Audio Processor chip, and an Audio Codec. All 5 chips in the set are implemented in a 0.5 or 0.6 micron CMOS process. Each of the chips implement digital signal processing algorithms of varying levels of complexity and flexibility. These levels range from standard interpolation and decimation filter implementations found on the Audio Codec to dual programmable digital signal processor cores found on the Bus Interface and Audio Processor chip. A top level description of the chip set architecture is presented, along with a functional description of a typical video teleconferencing system based on this chip set. This is followed by a top level description of the various digital signal processing architectures and approaches used in the individual chips in the chip set.

  9. Flip-chip packaging of piezoresistive barometric pressure sensors

    NASA Astrophysics Data System (ADS)

    Waber, T.; Pahl, W.; Schmidt, M.; Feiertag, G.; Stufler, S.; Dudek, R.; Leidl, A.

    2013-05-01

    To miniaturize piezoresistive barometric pressure sensors we have developed a package using flip-chip bonding. However, in a standard flip-chip package the different coefficients of thermal expansion (CTE) of chip and substrate and strong mechanical coupling by the solder bumps would lead to stress in the sensor chip which is not acceptable for piezoresistive pressure sensors. To overcome this problem we have developed a new ultra low stress flip-chip packaging technology. In this new packaging technology for pressure sensors first an under bump metallization (UBM) is patterned on the sensor wafer. As the next step solder bumps are deposited. After wafer-dicing the chips are flip-chip bonded on copper springs within a ceramic cavity. As sources of residual stress we identified the copper springs, the UBM and the solder bumps on the sensor chip. Different CTEs of the silicon chip and the UBM/solder lead to creep strain in the aluminum metallization between UBM and chip. As a consequence a temperature hysteresis can be measured.

  10. Managing DNA polymerases: coordinating DNA replication, DNA repair, and DNA recombination.

    PubMed

    Sutton, M D; Walker, G C

    2001-07-17

    Two important and timely questions with respect to DNA replication, DNA recombination, and DNA repair are: (i) what controls which DNA polymerase gains access to a particular primer-terminus, and (ii) what determines whether a DNA polymerase hands off its DNA substrate to either a different DNA polymerase or to a different protein(s) for the completion of the specific biological process? These questions have taken on added importance in light of the fact that the number of known template-dependent DNA polymerases in both eukaryotes and in prokaryotes has grown tremendously in the past two years. Most notably, the current list now includes a completely new family of enzymes that are capable of replicating imperfect DNA templates. This UmuC-DinB-Rad30-Rev1 superfamily of DNA polymerases has members in all three kingdoms of life. Members of this family have recently received a great deal of attention due to the roles they play in translesion DNA synthesis (TLS), the potentially mutagenic replication over DNA lesions that act as potent blocks to continued replication catalyzed by replicative DNA polymerases. Here, we have attempted to summarize our current understanding of the regulation of action of DNA polymerases with respect to their roles in DNA replication, TLS, DNA repair, DNA recombination, and cell cycle progression. In particular, we discuss these issues in the context of the Gram-negative bacterium, Escherichia coli, that contains a DNA polymerase (Pol V) known to participate in most, if not all, of these processes.

  11. DNA polymerases and cancer

    PubMed Central

    Lange, Sabine S.; Takata, Kei-ichi; Wood, Richard D.

    2013-01-01

    There are fifteen different DNA polymerases encoded in mammalian genomes, which are specialized for replication, repair or the tolerance of DNA damage. New evidence is emerging for lesion-specific and tissue-specific functions of DNA polymerases. Many point mutations that occur in cancer cells arise from the error-generating activities of DNA polymerases. However, the ability of some of these enzymes to bypass DNA damage may actually defend against chromosome instability in cells and at least one DNA polymerase, POLζ, is a suppressor of spontaneous tumorigenesis. Because DNA polymerases can help cancer cells tolerate DNA damage, some of these enzymes may be viable targets for therapeutic strategies. PMID:21258395

  12. DNA systematics. Volume II

    SciTech Connect

    Dutta, S.K.

    1986-01-01

    This book discusses the following topics: PLANTS: PLANT DNA: Contents and Systematics. Repeated DNA Sequences and Polyploidy in Cereal Crops. Homology of Nonrepeated DNA Sequences in Phylogeny of Fungal Species. Chloropast DNA and Phylogenetic Relationships. rDNA: Evolution Over a Billion Years. 23S rRNA-derived Small Ribosomal RNAs: Their Structure and Evolution with Reference to Plant Phylogeny. Molecular Analysis of Plant DNA Genomes: Conserved and Diverged DNA Sequences. A Critical Review of Some Terminologies Used for Additional DNA in Plant Chromosomes and Index.

  13. A Fully Integrated Paperfluidic Molecular Diagnostic Chip for the Extraction, Amplification, and Detection of Nucleic Acids from Clinical Samples

    PubMed Central

    Rodriguez, Natalia M.; Wong, Winnie S.; Liu, Lena; Dewar, Rajan; Klapperich, Catherine M.

    2016-01-01

    Paper diagnostics have successfully been employed to detect the presence of antigens or small molecules in clinical samples through immunoassays; however, the detection of many disease targets relies on the much higher sensitivity and specificity achieved via nucleic acid amplification tests (NAAT). The steps involved in NAAT have recently begun to be explored in paper matrices, and our group, among others, has reported on paper-based extraction, amplification, and detection of DNA and RNA targets. Here, we integrate these paper-based NAAT steps onto a single paperfluidic chip in a modular, foldable system that allows for fully integrated fluidic handling from sample to result. We showcase the functionality of the chip by combining nucleic acid isolation, isothermal amplification, and lateral flow detection of human papillomavirus (HPV) 16 DNA directly from crude cervical specimens in under 1 hour for rapid, early detection of cervical cancer. The chip is made entirely of paper and adhesive sheets, making it low-cost, portable, and disposable, and offering the potential for a point-of-care molecular diagnostic platform even in remote and resource-limited settings. PMID:26785636

  14. A Decade of Boon or Burden: What Has the CHIP Ever Done for Cellular Protein Quality Control Mechanism Implicated in Neurodegeneration and Aging?

    PubMed Central

    Joshi, Vibhuti; Amanullah, Ayeman; Upadhyay, Arun; Mishra, Ribhav; Kumar, Amit; Mishra, Amit

    2016-01-01

    Cells regularly synthesize new proteins to replace old and abnormal proteins for normal cellular functions. Two significant protein quality control pathways inside the cellular milieu are ubiquitin proteasome system (UPS) and autophagy. Autophagy is known for bulk clearance of cytoplasmic aggregated proteins, whereas the specificity of protein degradation by UPS comes from E3 ubiquitin ligases. Few E3 ubiquitin ligases, like C-terminus of Hsc70-interacting protein (CHIP) not only take part in protein quality control pathways, but also plays a key regulatory role in other cellular processes like signaling, development, DNA damage repair, immunity and aging. CHIP targets misfolded proteins for their degradation through proteasome, as well as autophagy; simultaneously, with the help of chaperones, it also regulates folding attempts for misfolded proteins. The broad range of CHIP substrates and their associations with multiple pathologies make it a key molecule to work upon and focus for future therapeutic interventions. E3 ubiquitin ligase CHIP interacts and degrades many protein inclusions formed in neurodegenerative diseases. The presence of CHIP at various nodes of cellular protein-protein interaction network presents this molecule as a potential candidate for further research. In this review, we have explored a wide range of functionality of CHIP inside cells by a detailed presentation of its co-chaperone, E3 and E4 enzyme like functions, with central focus on its protein quality control roles in neurodegenerative diseases. We have also raised many unexplored but expected fundamental questions regarding CHIP functions, which generate hopes for its future applications in research, as well as drug discovery. PMID:27757073

  15. Detection of cystic fibrosis mutations in a GeneChip{trademark} assay format

    SciTech Connect

    Miyada, C.G.; Cronin, M.T.; Kim, S.M.

    1994-09-01

    We are developing assays for the detection of cystic fibrosis mutations based on DNA hybridization. A DNA sample is amplified by PCR, labeled by incorporating a fluorescein-tagged dNTP, enzymatically treated to produce smaller fragments and hybridized to a series of short (13-16 bases) oligonucleotides synthesized on a glass surface via photolithography. The hybrids are detected by eqifluorescence and mutations are identified by the specific pattern of hybridization. In a GeneChip assay, the chip surface is composed of a series of subarrays, each being specific for a particular mutation. Each subarray is further subdivided into a series of probes (40 total), half based on the mutant sequence and the remainder based on the wild-type sequence. For each of the subarrays, there is a redundancy in the number of probes that should hybridize to either a wild-type or a mutant target. The multiple probe strategy provides sequence information for a short five base region overlapping the mutation site. In addition, homozygous wild-type and mutant as well as heterozygous samples are each identified by a specific pattern of hybridization. The small size of each probe feature (250 x 250 {mu}m{sup 2}) permits the inclusion of additional probes required to generate sequence information by hybridization.

  16. Aberrant DNA Methylation in Keratoacanthoma

    PubMed Central

    Nakagawa, Hidemi

    2016-01-01

    Background Keratoacanthoma (KA) is a self-limiting epidermal tumor for which histopathological examination sometimes suggests malignancy. Based on inconsistent clinical views, KA can be regarded as both a benign tumor and a variant of squamous cell carcinoma (SCC). Aberrant DNA methylation frequently occurs in malignant tumors but it scarcely occurs in benign tumors. Whether aberrant methylation occurs in KA has not been previously examined. Objective The aim is to elucidate whether aberrant methylation of CpG islands (CGI) containing a high density of cytosine-guanine dinucleotide (CpG) sites occurs in KA. Methods Five SCC cell lines, two cultured samples of normal human epidermal keratinocytes (NHEKs), 18 clinical SCC samples, and 21 clinical KA samples were analyzed with Infinium HumanMethylation450 BeadChips, quantitative real-time methylation-specific PCR (RT-MSP) and/or bisulfite sequencing. Results Genome-wide analyses of NHEK, KA, and SCC indicated that there was a greater number of aberrantly hypermethylated CGIs in SCC than in KA and there were aberrantly hypermethylated CGIs which are common in both. Among the common hypermethylated CGIs, RT-MSP and bisulfite sequencing targeting CGIs located on CCDC17, PVR, and MAP3K11 gene bodies also showed that methylation levels were significantly higher in KA than in normal epidermis. Statistical analyses suggested that the methylation level of CGI located on PVR in SCC might be correlated to lymph node metastasis (P = 0.013, Mann-Whitney U test) and that the methylation level of CGI in MAP3K11 in KA might be correlated to age (P = 0.031, linear regression analysis). Conclusion Aberrant DNA methylation occurs in KA. PMID:27788211

  17. Autonomous DNA computing machine based on photochemical gate transition.

    PubMed

    Ogasawara, Shinzi; Ami, Takehiro; Fujimoto, Kenzo

    2008-08-01

    We report the construction of a one-pot autonomous DNA computing machine based on photochemical gate transition (photocleavage, hybridization, and photoligation), and we performed binary digit additions using this machine. In our method, both photochemical DNA manipulations previously reported, photoligation via 5-carboxyvinyldeoxyuridene (cvU) containing ODN and photocleavage via carbazole-modified ODN, were employed. The binary digit additions were autonomously carried out by one-time irradiation at 366 nm in the single test tube. The fluorescence readout by the DNA chip was in good agreement with the correct answer of binary digit additions. We believe that this system is easily applicable to correlation analysis between SNPs as well as other binary digit processing, such as subtraction.

  18. DNA Nanotechnology-- Architectures Designed with DNA

    NASA Astrophysics Data System (ADS)

    Han, Dongran

    As the genetic information storage vehicle, deoxyribonucleic acid (DNA) molecules are essential to all known living organisms and many viruses. It is amazing that such a large amount of information about how life develops can be stored in these tiny molecules. Countless scientists, especially some biologists, are trying to decipher the genetic information stored in these captivating molecules. Meanwhile, another group of researchers, nanotechnologists in particular, have discovered that the unique and concise structural features of DNA together with its information coding ability can be utilized for nano-construction efforts. This idea culminated in the birth of the field of DNA nanotechnology which is the main topic of this dissertation. The ability of rationally designed DNA strands to self-assemble into arbitrary nanostructures without external direction is the basis of this field. A series of novel design principles for DNA nanotechnology are presented here, from topological DNA nanostructures to complex and curved DNA nanostructures, from pure DNA nanostructures to hybrid RNA/DNA nanostructures. As one of the most important and pioneering fields in controlling the assembly of materials (both DNA and other materials) at the nanoscale, DNA nanotechnology is developing at a dramatic speed and as more and more construction approaches are invented, exciting advances will emerge in ways that we may or may not predict.

  19. DNA vaccines: a simple DNA sensing matter?

    PubMed

    Coban, Cevayir; Kobiyama, Kouji; Jounai, Nao; Tozuka, Miyuki; Ishii, Ken J

    2013-10-01

    Since the introduction of DNA vaccines two decades ago, this attractive strategy has been hampered by its low immunogenicity in humans. Studies conducted to improve the immunogenicity of DNA vaccines have shown that understanding the mechanism of action of DNA vaccines might be the key to successfully improving their immunogenicity. Our current understanding is that DNA vaccines induce innate and adaptive immune responses in two ways: (1) encoded protein (or polypeptide) antigen(s) by the DNA plasmid can be expressed in stromal cells (i.e., muscle cells) as well as DCs, where these antigens are processed and presented to naïve CD4 or CD8 T cells either by direct or cross presentation, respectively; and (2) the transfected DNA plasmid itself may bind to an un-identified cytosolic DNA sensor and activate the TBK1-STING pathway and the production of type I interferons (IFNs) which function as an adjuvant. Recent studies investigating double-stranded cytosolic DNA sensor(s) have highlighted new mechanisms in which cytosolic DNA may release secondary metabolites, which are in turn recognized by a novel DNA sensing machinery. Here, we discuss these new metabolites and the possibilities of translating this knowledge into improved immunogenicity for DNA vaccines.

  20. Hot Chips and Hot Interconnects for High End Computing Systems

    NASA Technical Reports Server (NTRS)

    Saini, Subhash

    2005-01-01

    I will discuss several processors: 1. The Cray proprietary processor used in the Cray X1; 2. The IBM Power 3 and Power 4 used in an IBM SP 3 and IBM SP 4 systems; 3. The Intel Itanium and Xeon, used in the SGI Altix systems and clusters respectively; 4. IBM System-on-a-Chip used in IBM BlueGene/L; 5. HP Alpha EV68 processor used in DOE ASCI Q cluster; 6. SPARC64 V processor, which is used in the Fujitsu PRIMEPOWER HPC2500; 7. An NEC proprietary processor, which is used in NEC SX-6/7; 8. Power 4+ processor, which is used in Hitachi SR11000; 9. NEC proprietary processor, which is used in Earth Simulator. The IBM POWER5 and Red Storm Computing Systems will also be discussed. The architectures of these processors will first be presented, followed by interconnection networks and a description of high-end computer systems based on these processors and networks. The performance of various hardware/programming model combinations will then be compared, based on latest NAS Parallel Benchmark results (MPI, OpenMP/HPF and hybrid (MPI + OpenMP). The tutorial will conclude with a discussion of general trends in the field of high performance computing, (quantum computing, DNA computing, cellular engineering, and neural networks).

  1. Flip-chip light emitting diode with resonant optical microcavity

    DOEpatents

    Gee, James M.; Bogart, Katherine H.A.; Fischer, Arthur J.

    2005-11-29

    A flip-chip light emitting diode with enhanced efficiency. The device structure employs a microcavity structure in a flip-chip configuration. The microcavity enhances the light emission in vertical modes, which are readily extracted from the device. Most of the rest of the light is emitted into waveguided lateral modes. Flip-chip configuration is advantageous for light emitting diodes (LEDs) grown on dielectric substrates (e.g., gallium nitride LEDs grown on sapphire substrates) in general due to better thermal dissipation and lower series resistance. Flip-chip configuration is advantageous for microcavity LEDs in particular because (a) one of the reflectors is a high-reflectivity metal ohmic contact that is already part of the flip-chip configuration, and (b) current conduction is only required through a single distributed Bragg reflector. Some of the waveguided lateral modes can also be extracted with angled sidewalls used for the interdigitated contacts in the flip-chip configuration.

  2. Fungi on fuel wood chips in a home

    SciTech Connect

    Miller, J.D.; Schneider, M.H.; Whitney, N.J.

    1982-01-01

    Softwood tops and branch fuel chips with high moisture contents were subject to biological heating in storage. This was due primarily to infestations of mesophilic (ca. 5 X 10 to the power of 4 propagules/g dry weight wood) and thermophilic (ca. 1.6 X 10 to the power of 6 propagules/g dry weight wood) fungi. Loading chips into a home fuel-chip furnace resulted in the distribution of fungal propagules throughout the basement and upper floors. Many of the species isolated are human allergens and pathogens. The results suggest that dry storage of chips (that is, environmental conditions which do not allow fungal growth) is important to avoid propagation of allergenic and pathogenic fungi. They also suggest that chips which have been subject to biological heating should not be transported into a home without precautions. Individuals handling chips should wear dust masks, and take other measures to avoid prolonged contact and contamination of living quarters. (Refs. 10).

  3. Processing and quality evaluation of sweet potato chips.

    PubMed

    Akpapunam, M A; Abiante, D A

    1991-10-01

    A study was conducted to develop a process for producing sweet potato chips. Sweet potato tubers sliced to 0.5 by 0.5 cm size were dehydrated at 70 degrees C for various times (0, 90, 105, 120, 135, 150, 165 min). Determination of the moisture content of the dehydrated chips and sensory evaluation of the dehydrated and fried chips were carried out to establish optimum dehydration time and moisture which corresponded to optimum quality. Blanching the slices in water and 1% sodium metabisulfite solution respectively prior to the dehydration significantly (P greater than 0.05) improved the color and general acceptability of the chips over those immersed in water. The process development resulted in about 26 to 76% decrease in the ascorbic acid content of the chips. Significant changes also occurred in the total and reducing sugars of the chip following partial dehydration.

  4. A primary battery-on-a-chip using monolayer graphene

    NASA Astrophysics Data System (ADS)

    Iost, Rodrigo M.; Crespilho, Frank N.; Kern, Klaus; Balasubramanian, Kannan

    2016-07-01

    We present here a bottom-up approach for realizing on-chip on-demand batteries starting out with chemical vapor deposition-grown graphene. Single graphene monolayers contacted by electrode lines on a silicon chip serve as electrodes. The anode and cathode are realized by electrodeposition of zinc and copper respectively onto graphene, leading to the realization of a miniature graphene-based Daniell cell on a chip. The electrolyte is housed partly in a gel and partly in liquid form in an on-chip enclosure molded using a 3d printer or made out of poly(dimethylsiloxane). The realized batteries provide a stable voltage (∼1.1 V) for many hours and exhibit capacities as high as 15 μAh, providing enough power to operate a pocket calculator. The realized batteries show promise for deployment as on-chip power sources for autonomous systems in lab-on-a-chip or biomedical applications.

  5. Neural network chips for trigger purposes in high energy physics

    SciTech Connect

    Gemmeke, H.; Eppler, W.; Fischer, T.

    1996-12-31

    Two novel neural chips SAND (Simple Applicable Neural Device) and SIOP (Serial Input - Operating Parallel) are described. Both are highly usable for hardware triggers in particle physics. The chips are optimized for a high input data rate at a very low cost basis. The performance of a single SAND chip is 200 MOPS due to four parallel 16 bit multipliers and 40 bit adders working in one clock cycle. The chip is able to implement feedforward neural networks, Kohonen feature maps and radial basis function networks. Four chips will be implemented on a PCI-board for simulation and on a VUE board for trigger and on- and off-line analysis. For small sized feedforward neural networks the bit-serial neuro-chip SIOP may lead to even smaller latencies because each synaptic connection is implemented by its own bit serial multiplier and adder.

  6. On-chip particle trapping and manipulation

    NASA Astrophysics Data System (ADS)

    Leake, Kaelyn Danielle

    The ability to control and manipulate the world around us is human nature. Humans and our ancestors have used tools for millions of years. Only in recent years have we been able to control objects at such small levels. In order to understand the world around us it is frequently necessary to interact with the biological world. Optical trapping and manipulation offer a non-invasive way to move, sort and interact with particles and cells to see how they react to the world around them. Optical tweezers are ideal in their abilities but they require large, non-portable, and expensive setups limiting how and where we can use them. A cheap portable platform is required in order to have optical manipulation reach its full potential. On-chip technology offers a great solution to this challenge. We focused on the Liquid-Core Anti-Resonant Reflecting Optical Waveguide (liquid-core ARROW) for our work. The ARROW is an ideal platform, which has anti-resonant layers which allow light to be guided in liquids, allowing for particles to easily be manipulated. It is manufactured using standard silicon manufacturing techniques making it easy to produce. The planner design makes it easy to integrate with other technologies. Initially I worked to improve the ARROW chip by reducing the intersection losses and by reducing the fluorescence and background on the ARROW chip. The ARROW chip has already been used to trap and push particles along its channel but here I introduce several new methods of particle trapping and manipulation on the ARROW chip. Traditional two beam traps use two counter propagating beams. A trapping scheme that uses two orthogonal beams which counter to first instinct allow for trapping at their intersection is introduced. This scheme is thoroughly predicted and analyzed using realistic conditions. Simulations of this method were done using a program which looks at both the fluidics and optical sources to model complex situations. These simulations were also used to

  7. Sub-micro-liter Electrochemical Single-Nucleotide-Polymorphism Detector for Lab-on-a-Chip System

    NASA Astrophysics Data System (ADS)

    Tanaka, Hiroyuki; Fiorini, Paolo; Peeters, Sara; Majeed, Bivragh; Sterken, Tom; de Beeck, Maaike Op; Hayashi, Miho; Yaku, Hidenobu; Yamashita, Ichiro

    2012-04-01

    A sub-micro-liter single-nucleotide-polymorphism (SNP) detector for lab-on-a-chip applications is developed. This detector enables a fast, sensitive, and selective SNP detection directly from human blood. The detector is fabricated on a Si substrate by a standard complementary metal oxide semiconductor/micro electro mechanical systems (CMOS/MEMS) process and Polydimethylsiloxane (PDMS) molding. Stable and reproducible measurements are obtained by implementing an on-chip Ag/AgCl electrode and encapsulating the detector. The detector senses the presence of SNPs by measuring the concentration of pyrophosphoric acid generated during selective DNA amplification. A 0.5-µL-volume detector enabled the successful performance of the typing of a SNP within the ABO gene using human blood. The measured sensitivity is 566 pA/µM.

  8. Technology for melting amber chips to produce a solid block

    NASA Astrophysics Data System (ADS)

    Vikhareva, A. S.; Melnikov, A. G.; Utyev, O. M.

    2016-04-01

    This research is relevant, because the bulk of the mined amber comes in amber chips. Therefore, we have decided to review the current ways of melting amber chips to develop the most technologically efficient algorithm and to use it further for producing decorative items. The purpose of the work is to perfect the technology of obtaining whole-piece amber from amber chips and to explore the usability of the obtained material in decorative items and jewelry.

  9. Lithographic chip identification: meeting the failure analysis challenge

    NASA Astrophysics Data System (ADS)

    Perkins, Lynn; Riddell, Kevin G.; Flack, Warren W.

    1992-06-01

    This paper describes a novel method using stepper photolithography to uniquely identify individual chips for permanent traceability. A commercially available 1X stepper is used to mark chips with an identifier or `serial number' which can be encoded with relevant information for the integrated circuit manufacturer. The permanent identification of individual chips can improve current methods of quality control, failure analysis, and inventory control. The need for this technology is escalating as manufacturers seek to provide six sigma quality control for their products and trace fabrication problems to their source. This need is especially acute for parts that fail after packaging and are returned to the manufacturer for analysis. Using this novel approach, failure analysis data can be tied back to a particular batch, wafer, or even a position within a wafer. Process control can be enhanced by identifying the root cause of chip failures. Chip identification also addresses manufacturers concerns with increasing incidences of chip theft. Since chips currently carry no identification other than the manufacturer's name and part number, recovery efforts are hampered by the inability to determine the sales history of a specific packaged chip. A definitive identifier or serial number for each chip would address this concern. The results of chip identification (patent pending) are easily viewed through a low power microscope. Batch number, wafer number, exposure step, and chip location within the exposure step can be recorded, as can dates and other items of interest. An explanation of the chip identification procedure and processing requirements are described. Experimental testing and results are presented, and potential applications are discussed.

  10. Quantitative DNA fiber mapping

    DOEpatents

    Gray, Joe W.; Weier, Heinz-Ulrich G.

    1998-01-01

    The present invention relates generally to the DNA mapping and sequencing technologies. In particular, the present invention provides enhanced methods and compositions for the physical mapping and positional cloning of genomic DNA. The present invention also provides a useful analytical technique to directly map cloned DNA sequences onto individual stretched DNA molecules.

  11. Low-power chip-level optical interconnects based on bulk-silicon single-chip photonic transceivers

    NASA Astrophysics Data System (ADS)

    Kim, Gyungock; Park, Hyundai; Joo, Jiho; Jang, Ki-Seok; Kwack, Myung-Joon; Kim, Sanghoon; Kim, In Gyoo; Kim, Sun Ae; Oh, Jin Hyuk; Park, Jaegyu; Kim, Sanggi

    2016-03-01

    We present new scheme for chip-level photonic I/Os, based on monolithically integrated vertical photonic devices on bulk silicon, which increases the integration level of PICs to a complete photonic transceiver (TRx) including chip-level light source. A prototype of the single-chip photonic TRx based on a bulk silicon substrate demonstrated 20 Gb/s low power chip-level optical interconnects between fabricated chips, proving that this scheme can offer compact low-cost chip-level I/O solutions and have a significant impact on practical electronic-photonic integration in high performance computers (HPC), cpu-memory interface, 3D-IC, and LAN/SAN/data-center and network applications.

  12. Viral diagnosis in Indian livestock using customized microarray chips.

    PubMed

    Yadav, Brijesh S; Pokhriyal, Mayank; Ratta, Barkha; Kumar, Ajay; Saxena, Meeta; Sharma, Bhaskar

    2015-01-01

    Viral diagnosis in Indian livestock using customized microarray chips is gaining momentum in recent years. Hence, it is possible to design customized microarray chip for viruses infecting livestock in India. Customized microarray chips identified Bovine herpes virus-1 (BHV-1), Canine Adeno Virus-1 (CAV-1), and Canine Parvo Virus-2 (CPV-2) in clinical samples. Microarray identified specific probes were further confirmed using RT-PCR in all clinical and known samples. Therefore, the application of microarray chips during viral disease outbreaks in Indian livestock is possible where conventional methods are unsuitable. It should be noted that customized application requires a detailed cost efficiency calculation.

  13. Alignment of microcircuit chips using optically smeared images.

    PubMed

    Lewis, R W

    1979-02-01

    An optical method for determining the position of microcircuit chips for wirebonding or electrical testing stations was evaluated. Optically smearing the chip image in one direction with a cylindrical lens produces a convenient means for determining both chip angular orientation and position. Digitized images from a linear photodiode array camera were analyzed. The results show that a class of microcircuit chips with medium scale integration can be aligned in angle and position to a higher accuracy than required for wirebonding and electrical testing stations. PMID:20208714

  14. Real time image processing with an analog vision chip system.

    PubMed

    Kameda, S; Honda, A; Yagi, T

    1999-10-01

    A linear analog network model is proposed to characterize the function of the outer retinal circuit in terms of the standard regularization theory. Inspired by the function and the architecture of the model, a vision chip has been designed using analog CMOS Very Large Scale Integrated circuit technology. In the chip, sample/hold amplifier circuits are incorporated to compensate for statistic transistor mismatches. Accordingly, extremely low noise outputs were obtained from the chip. Using the chip and a zero-crossing detector, edges of given images were effectively extracted in indoor illumination.

  15. A lab-on-a-chip device for rapid identification of avian influenza viral RNA by solid-phase PCR.

    PubMed

    Sun, Yi; Dhumpa, Raghuram; Bang, Dang Duong; Høgberg, Jonas; Handberg, Kurt; Wolff, Anders

    2011-04-21

    The endemic of Avian Influenza Virus (AIV) in Asia and epizootics in some European regions have caused serious economic losses. Multiplex reverse-transcriptase (RT) PCR has been developed to detect and subtype AIV. However, the number of targets that can be amplified in a single run is limited because of uncontrollable primer-primer interferences. In this paper, we describe a lab-on-a-chip device for fast AIV screening by integrating DNA microarray-based solid-phase PCR on a microfluidic chip. A simple UV cross-linking method was used to immobilize the DNA probes on unmodified glass surface, which makes it convenient to integrate microarray with microfluidics. This solid-phase RT-PCR method combined RT amplification of extracted RNA in the liquid phase and species-specific nested PCR on the solid phase. Using the developed approach, AIV viruses and their subtypes were unambiguously identified by the distinct patterns of amplification products. The whole process was reduced to less than 1 hour and the sample volume used in the microfluidic chip was at least 10 times less than in the literature. By spatially separating the primers, highly multiplexed amplification can be performed in solid-phase PCR. Moreover, multiplex PCR and sequence detection were done in one step, which greatly simplified the assay and reduced the processing time. Furthermore, by incorporating the microarray into a microchamber-based PCR chip, the sample and the reagent consumption were greatly reduced, and the problems of bubble formation and solution evaporation were effectively prevented. This microarray-based PCR microchip can be widely employed for virus detection and effective surveillance in wild avian and in poultry productions.

  16. On-chip plasmonic waveguide optical waveplate

    NASA Astrophysics Data System (ADS)

    Gao, Linfei; Huo, Yijie; Zang, Kai; Paik, Seonghyun; Chen, Yusi; Harris, James S.; Zhou, Zhiping

    2015-10-01

    Polarization manipulation is essential in almost every photonic system ranging from telecommunications to bio-sensing to quantum information. This is traditionally achieved using bulk waveplates. With the developing trend of photonic systems towards integration and miniaturization, the need for an on-chip waveguide type waveplate becomes extremely urgent. However, this is very challenging using conventional dielectric waveguides, which usually require complex 3D geometries to alter the waveguide symmetry and are also difficult to create an arbitrary optical axis. Recently, a waveguide waveplate was realized using femtosecond laser writing, but the device length is in millimeter range. Here, for the first time we propose and experimentally demonstrate an ultracompact, on-chip waveplate using an asymmetric hybrid plasmonic waveguide to create an arbitrary optical axis. The device is only in several microns length and produced in a flexible integratable IC compatible format, thus opening up the potential for integration into a broad range of systems.

  17. On-chip plasmonic waveguide optical waveplate

    PubMed Central

    Gao, Linfei; Huo, Yijie; Zang, Kai; Paik, Seonghyun; Chen, Yusi; Harris, James S.; Zhou, Zhiping

    2015-01-01

    Polarization manipulation is essential in almost every photonic system ranging from telecommunications to bio-sensing to quantum information. This is traditionally achieved using bulk waveplates. With the developing trend of photonic systems towards integration and miniaturization, the need for an on-chip waveguide type waveplate becomes extremely urgent. However, this is very challenging using conventional dielectric waveguides, which usually require complex 3D geometries to alter the waveguide symmetry and are also difficult to create an arbitrary optical axis. Recently, a waveguide waveplate was realized using femtosecond laser writing, but the device length is in millimeter range. Here, for the first time we propose and experimentally demonstrate an ultracompact, on-chip waveplate using an asymmetric hybrid plasmonic waveguide to create an arbitrary optical axis. The device is only in several microns length and produced in a flexible integratable IC compatible format, thus opening up the potential for integration into a broad range of systems. PMID:26507563

  18. Invisibility Cloak Printed on a Photonic Chip.

    PubMed

    Feng, Zhen; Wu, Bing-Hong; Zhao, Yu-Xi; Gao, Jun; Qiao, Lu-Feng; Yang, Ai-Lin; Lin, Xiao-Feng; Jin, Xian-Min

    2016-01-01

    Invisibility cloak capable of hiding an object can be achieved by properly manipulating electromagnetic field. Such a remarkable ability has been shown in transformation and ray optics. Alternatively, it may be realistic to create a spatial cloak by means of confining electromagnetic field in three-dimensional arrayed waveguides and introducing appropriate collective curvature surrounding an object. We realize the artificial structure in borosilicate by femtosecond laser direct writing, where we prototype up to 5,000 waveguides to conceal millimeter-scale volume. We characterize the performance of the cloak by normalized cross correlation, tomography analysis and continuous three-dimensional viewing angle scan. Our results show invisibility cloak can be achieved in waveguide optics. Furthermore, directly printed invisibility cloak on a photonic chip may enable controllable study and novel applications in classical and quantum integrated photonics, such as invisualising a coupling or swapping operation with on-chip circuits of their own.

  19. On-chip plasmonic waveguide optical waveplate.

    PubMed

    Gao, Linfei; Huo, Yijie; Zang, Kai; Paik, Seonghyun; Chen, Yusi; Harris, James S; Zhou, Zhiping

    2015-01-01

    Polarization manipulation is essential in almost every photonic system ranging from telecommunications to bio-sensing to quantum information. This is traditionally achieved using bulk waveplates. With the developing trend of photonic systems towards integration and miniaturization, the need for an on-chip waveguide type waveplate becomes extremely urgent. However, this is very challenging using conventional dielectric waveguides, which usually require complex 3D geometries to alter the waveguide symmetry and are also difficult to create an arbitrary optical axis. Recently, a waveguide waveplate was realized using femtosecond laser writing, but the device length is in millimeter range. Here, for the first time we propose and experimentally demonstrate an ultracompact, on-chip waveplate using an asymmetric hybrid plasmonic waveguide to create an arbitrary optical axis. The device is only in several microns length and produced in a flexible integratable IC compatible format, thus opening up the potential for integration into a broad range of systems.

  20. Invisibility Cloak Printed on a Photonic Chip

    PubMed Central

    Feng, Zhen; Wu, Bing-Hong; Zhao, Yu-Xi; Gao, Jun; Qiao, Lu-Feng; Yang, Ai-Lin; Lin, Xiao-Feng; Jin, Xian-Min

    2016-01-01

    Invisibility cloak capable of hiding an object can be achieved by properly manipulating electromagnetic field. Such a remarkable ability has been shown in transformation and ray optics. Alternatively, it may be realistic to create a spatial cloak by means of confining electromagnetic field in three-dimensional arrayed waveguides and introducing appropriate collective curvature surrounding an object. We realize the artificial structure in borosilicate by femtosecond laser direct writing, where we prototype up to 5,000 waveguides to conceal millimeter-scale volume. We characterize the performance of the cloak by normalized cross correlation, tomography analysis and continuous three-dimensional viewing angle scan. Our results show invisibility cloak can be achieved in waveguide optics. Furthermore, directly printed invisibility cloak on a photonic chip may enable controllable study and novel applications in classical and quantum integrated photonics, such as invisualising a coupling or swapping operation with on-chip circuits of their own. PMID:27329510

  1. Microengineered physiological biomimicry: organs-on-chips.

    PubMed

    Huh, Dongeun; Torisawa, Yu-suke; Hamilton, Geraldine A; Kim, Hyun Jung; Ingber, Donald E

    2012-06-21

    Microscale engineering technologies provide unprecedented opportunities to create cell culture microenvironments that go beyond current three-dimensional in vitro models by recapitulating the critical tissue-tissue interfaces, spatiotemporal chemical gradients, and dynamic mechanical microenvironments of living organs. Here we review recent advances in this field made over the past two years that are focused on the development of 'Organs-on-Chips' in which living cells are cultured within microfluidic devices that have been microengineered to reconstitute tissue arrangements observed in living organs in order to study physiology in an organ-specific context and to develop specialized in vitro disease models. We discuss the potential of organs-on-chips as alternatives to conventional cell culture models and animal testing for pharmaceutical and toxicology applications. We also explore challenges that lie ahead if this field is to fulfil its promise to transform the future of drug development and chemical safety testing.

  2. Ion trap in a semiconductor chip

    NASA Astrophysics Data System (ADS)

    Stick, D.; Hensinger, W. K.; Olmschenk, S.; Madsen, M. J.; Schwab, K.; Monroe, C.

    2006-01-01

    The electromagnetic manipulation of isolated atoms has led to many advances in physics, from laser cooling and Bose-Einstein condensation of cold gases to the precise quantum control of individual atomic ions. Work on miniaturizing electromagnetic traps to the micrometre scale promises even higher levels of control and reliability. Compared with `chip traps' for confining neutral atoms, ion traps with similar dimensions and power dissipation offer much higher confinement forces and allow unparalleled control at the single-atom level. Moreover, ion microtraps are of great interest in the development of miniature mass-spectrometer arrays, compact atomic clocks and, most notably, large-scale quantum information processors. Here we report the operation of a micrometre-scale ion trap, fabricated on a monolithic chip using semiconductor micro-electromechanical systems (MEMS) technology. We confine, laser cool and measure heating of a single 111Cd+ ion in an integrated radiofrequency trap etched from a doped gallium-arsenide heterostructure.

  3. Invisibility Cloak Printed on a Photonic Chip

    NASA Astrophysics Data System (ADS)

    Feng, Zhen; Wu, Bing-Hong; Zhao, Yu-Xi; Gao, Jun; Qiao, Lu-Feng; Yang, Ai-Lin; Lin, Xiao-Feng; Jin, Xian-Min

    2016-06-01

    Invisibility cloak capable of hiding an object can be achieved by properly manipulating electromagnetic field. Such a remarkable ability has been shown in transformation and ray optics. Alternatively, it may be realistic to create a spatial cloak by means of confining electromagnetic field in three-dimensional arrayed waveguides and introducing appropriate collective curvature surrounding an object. We realize the artificial structure in borosilicate by femtosecond laser direct writing, where we prototype up to 5,000 waveguides to conceal millimeter-scale volume. We characterize the performance of the cloak by normalized cross correlation, tomography analysis and continuous three-dimensional viewing angle scan. Our results show invisibility cloak can be achieved in waveguide optics. Furthermore, directly printed invisibility cloak on a photonic chip may enable controllable study and novel applications in classical and quantum integrated photonics, such as invisualising a coupling or swapping operation with on-chip circuits of their own.

  4. The single-chip FASTBUS Slave Interface

    SciTech Connect

    Nelson, R.O.; Machen, D.R.; Downing, R.W.

    1990-12-31

    A single-chip implementation of the general-purpose FASTBUS Slave Interface (FSI) has been developed in ECL gate-array technology. The FSI will occupy only 1.6% of the available circuit board space while providing a complete 32-bit interface to the FASTBUS. All mandatory slave-interface requirements of IEEE 960 are supported, in addition to several non-mandatory requirements and the optional, extended MS code features. Geographic, logical, and broadcast addressing are implemented using on-chip registers. An optional multiple-module addressing technique is included that allows participating modules residing on a common crate or cable segment to respond as if individually addressed in sequence. The user interface provided by the FSI allows control of slave status-response and connection timing for both address and data cycles. The BIT1 ECL array technology used for the FSI allows direct connections to the FASTBUS, eliminating the need for external driver/receiver buffers.

  5. Fault injection via on-chip debugging in the internal memory of systems-on-chip processor

    NASA Astrophysics Data System (ADS)

    Chekmarev, S. A.; Khanov, V. Kh

    2015-10-01

    The paper presents an on-chip debugging method for the injection of single faults in the processor cores of systems-on-chip. The method consists in the placement of faults injection infrastructure in a system-on-chip as an intellectual property core. This simplifies the fault injection environment, reduces delays injection and improves the performance, as well as allows doing long autonomous campaign for injection of faults without the use of external devices.

  6. Flip-chip and backside techniques.

    SciTech Connect

    Bernhard-Hofer, Karoline; Barton, Daniel Lee; Cole, Edward Isaac, Jr.

    2010-08-01

    State-of-the-art techniques for failure localization and design modification through bulk silicon are essential for multi-level metallization and new, flip chip packaging methods. The tutorial reviews the transmission of light through silicon, sample preparation, and backside defect localization techniques that are both currently available and under development. The techniques covered include emission microscopy, scanning laser microscope based techniques (electrooptic techniques, LIVA and its derivatives), and other non-IR based tools (FIB, e-beam techniques, etc.).

  7. Pyramidal micromirrors for microsystems and atom chips

    NASA Astrophysics Data System (ADS)

    Trupke, M.; Ramirez-Martinez, F.; Curtis, E. A.; Ashmore, J. P.; Eriksson, S.; Hinds, E. A.; Moktadir, Z.; Gollasch, C.; Kraft, M.; Vijaya Prakash, G.; Baumberg, J. J.

    2006-02-01

    Concave pyramids are created in the (100) surface of a silicon wafer by anisotropic etching in potassium hydroxide. High quality micromirrors are then formed by sputtering gold onto the smooth silicon (111) faces of the pyramids. These mirrors show great promise as high quality optical devices suitable for integration into micro-optoelectromechanical systems and atom chips. We have shown that structures of this shape can be used to laser-cool and hold atoms in a magneto-optical trap.

  8. An all-solid-state, WDM silicon photonic digital link for chip-to-chip communications.

    PubMed

    Thacker, Hiren D; Zheng, Xuezhe; Lexau, Jon; Shafiiha, Roshanak; Shubin, Ivan; Lin, Shiyun; Djordjevic, Stevan; Amberg, Philip; Chang, Eric; Liu, Frankie; Simons, John; Lee, Jin-Hyoung; Abed, Arin; Liang, Hong; Luo, Ying; Yao, Jin; Feng, Dazeng; Asghari, Mehdi; Ho, Ron; Raj, Kannan; Cunningham, John E; Krishnamoorthy, Ashok V

    2015-05-18

    We describe a multiwavelength hybrid-integrated solid-state link on a 3 µm silicon-on-insulator (SOI) nanophotonic platform. The link spans three chips and employs germanium-silicon electroabsorption waveguide modulators, silicon transport waveguides, echelle gratings for multiplexing and demultiplexing, and pure germanium waveguide photo-detectors. The 8λ WDM Tx and Rx components are interconnected via a routing "bridge" chip using edge-coupled optical proximity communication. The packaged, retimed digital WDM link is demonstrated at 10 Gb/s and 10(-12) BER, with three wavelength channels consuming an on-chip power below 1.5 pJ/bit, excluding the external laser power.

  9. Integration of optoelectronic technologies for chip-to- chip interconnections and parallel pipeline processing

    NASA Astrophysics Data System (ADS)

    Wu, Jenming

    Digital information services such as multimedia systems and data communications require the processing and transfer of tremendous amount of data. These data need to be stored, accessed and delivered efficiently and reliably at high speed for various user applications. This represents a great challenge for current electronic systems. Electronics is effective in providing high performance processing and computation, but its input/outputs (I/Os) bandwidth is unable to scale with its processing power. The signal I/Os or interconnections are needed between processors and input devices, between processors for multiprocessor systems, and between processors and storage devices. Novel chip-to-chip interconnect technologies are needed to meet this challenge. This work integrates optoelectronic technologies for chip-to-chip interconnects and parallel pipeline processing. Photonic and electronic technologies are complementary to each other in the sense that electronics is more suitable for high-speed, low cost computation, and photonics is more suitable for high-bandwidth information transmission. Smart pixel technology uses electronics for logic switching and optics for chip-to- chip interconnects, thus combining the abilities of photonics and electronics nicely. This work describes both vertical and horizontal integration of smart pixel technologies for chip-to-chip optical interconnects and its applications. We present smart pixel VLSI designs in both hybrid CMOS/MQW smart pixel and monolithic GaAs smart pixel technologies. We use the CMOS/MQW technology for smart pixel array cellular logic (SPARCL) processors for SIMD parallel pipeline processing. We have tested the chip and constructed a prototype system for device characterization and system demonstration. We have verified the functionality of the system and characterized the electrical functions of the chip and the optoelectronic properties of the MQW devices. We have developed algorithms that utilize SPARCL for various

  10. Chip breaking system for automated machine tool

    DOEpatents

    Arehart, Theodore A.; Carey, Donald O.

    1987-01-01

    The invention is a rotary selectively directional valve assembly for use in an automated turret lathe for directing a stream of high pressure liquid machining coolant to the interface of a machine tool and workpiece for breaking up ribbon-shaped chips during the formation thereof so as to inhibit scratching or other marring of the machined surfaces by these ribbon-shaped chips. The valve assembly is provided by a manifold arrangement having a plurality of circumferentially spaced apart ports each coupled to a machine tool. The manifold is rotatable with the turret when the turret is positioned for alignment of a machine tool in a machining relationship with the workpiece. The manifold is connected to a non-rotational header having a single passageway therethrough which conveys the high pressure coolant to only the port in the manifold which is in registry with the tool disposed in a working relationship with the workpiece. To position the machine tools the turret is rotated and one of the tools is placed in a material-removing relationship of the workpiece. The passageway in the header and one of the ports in the manifold arrangement are then automatically aligned to supply the machining coolant to the machine tool workpiece interface for breaking up of the chips as well as cooling the tool and workpiece during the machining operation.

  11. Patch-clamp amplifiers on a chip.

    PubMed

    Weerakoon, Pujitha; Culurciello, Eugenio; Yang, Youshan; Santos-Sacchi, Joseph; Kindlmann, Peter J; Sigworth, Fred J

    2010-10-15

    We present the first, fully integrated, two-channel implementation of a patch-clamp measurement system. With this "PatchChip" two simultaneous whole-cell recordings can be obtained with rms noise of 8pA in a 10kHz bandwidth. The capacitance and series-resistance of the electrode can be compensated up to 10pF and 100MΩ respectively under computer control. Recordings of hERG and Na(v) 1.7 currents demonstrate the system's capabilities, which are on par with large, commercial patch-clamp instrumentation. By reducing patch-clamp amplifiers to a millimeter size micro-chip, this work paves the way to the realization of massively parallel, high-throughput patch-clamp systems for drug screening and ion-channel research. The PatchChip is implemented in a 0.5μm silicon-on-sapphire process; its size is 3×3mm(2) and the power consumption is 5mW per channel with a 3.3V power supply.

  12. Scanning magnetoresistance microscopy of atom chips.

    PubMed

    Volk, M; Whitlock, S; Wolff, C H; Hall, B V; Sidorov, A I

    2008-02-01

    Surface based geometries of microfabricated wires or patterned magnetic films can be used to magnetically trap and manipulate ultracold neutral atoms or Bose-Einstein condensates. We investigate the magnetic properties of such atom chips using a scanning magnetoresistive (MR) microscope with high spatial resolution and high field sensitivity. By comparing MR scans of a permanent magnetic atom chip to field profiles obtained using ultracold atoms, we show that MR sensors are ideally suited to observe small variations of the magnetic field caused by imperfections in the wires or magnetic materials which ultimately lead to fragmentation of ultracold atom clouds. Measurements are also provided for the magnetic field produced by a thin current-carrying wire with small geometric modulations along the edge. Comparisons of our measurements with a full numeric calculation of the current flow in the wire and the subsequent magnetic field show excellent agreement. Our results highlight the use of scanning MR microscopy as a convenient and powerful technique for precisely characterizing the magnetic fields produced near the surface of atom chips.

  13. Scalable NMR spectroscopy with semiconductor chips.

    PubMed

    Ha, Dongwan; Paulsen, Jeffrey; Sun, Nan; Song, Yi-Qiao; Ham, Donhee

    2014-08-19

    State-of-the-art NMR spectrometers using superconducting magnets have enabled, with their ultrafine spectral resolution, the determination of the structure of large molecules such as proteins, which is one of the most profound applications of modern NMR spectroscopy. Many chemical and biotechnological applications, however, involve only small-to-medium size molecules, for which the ultrafine resolution of the bulky, expensive, and high-maintenance NMR spectrometers is not required. For these applications, there is a critical need for portable, affordable, and low-maintenance NMR spectrometers to enable in-field, on-demand, or online applications (e.g., quality control, chemical reaction monitoring) and co-use of NMR with other analytical methods (e.g., chromatography, electrophoresis). As a critical step toward NMR spectrometer miniaturization, small permanent magnets with high field homogeneity have been developed. In contrast, NMR spectrometer electronics capable of modern multidimensional spectroscopy have thus far remained bulky. Complementing the magnet miniaturization, here we integrate the NMR spectrometer electronics into 4-mm(2) silicon chips. Furthermore, we perform various multidimensional NMR spectroscopies by operating these spectrometer electronics chips together with a compact permanent magnet. This combination of the spectrometer-electronics-on-a-chip with a permanent magnet represents a useful step toward miniaturization of the overall NMR spectrometer into a portable platform. PMID:25092330

  14. The easy road to genome-wide medium density SNP screening in a non-model species: development and application of a 10 K SNP-chip for the house sparrow (Passer domesticus).

    PubMed

    Hagen, Ingerid J; Billing, Anna M; Rønning, Bernt; Pedersen, Sindre A; Pärn, Henrik; Slate, Jon; Jensen, Henrik

    2013-05-01

    With the advent of next generation sequencing, new avenues have opened to study genomics in wild populations of non-model species. Here, we describe a successful approach to a genome-wide medium density Single Nucleotide Polymorphism (SNP) panel in a non-model species, the house sparrow (Passer domesticus), through the development of a 10 K Illumina iSelect HD BeadChip. Genomic DNA and cDNA derived from six individuals were sequenced on a 454 GS FLX system and generated a total of 1.2 million sequences, in which SNPs were detected. As no reference genome exists for the house sparrow, we used the zebra finch (Taeniopygia guttata) reference genome to determine the most likely position of each SNP. The 10 000 SNPs on the SNP-chip were selected to be distributed evenly across 31 chromosomes, giving on average one SNP per 100 000 bp. The SNP-chip was screened across 1968 individual house sparrows from four island populations. Of the original 10 000 SNPs, 7413 were found to be variable, and 99% of these SNPs were successfully called in at least 93% of all individuals. We used the SNP-chip to demonstrate the ability of such genome-wide marker data to detect population sub-division, and compared these results to similar analyses using microsatellites. The SNP-chip will be used to map Quantitative Trait Loci (QTL) for fitness-related phenotypic traits in natural populations.

  15. Evaluation of Pressure Stable Chip-to-Tube Fittings Enabling High-Speed Chip-HPLC with Mass Spectrometric Detection.

    PubMed

    Lotter, Carsten; Heiland, Josef J; Stein, Volkmar; Klimkait, Michael; Queisser, Marco; Belder, Detlev

    2016-08-01

    Appropriate chip-to-tube interfacing is an enabling technology for high-pressure and high-speed liquid chromatography on chip. For this purpose, various approaches, to connect pressure resistant glass chips with HPLC pumps working at pressures of up to 500 bar, were examined. Three side-port and one top-port connection approach were evaluated with regard to pressure stability and extra column band broadening. A clamp-based top-port approach enabled chip-HPLC-MS analysis of herbicides at the highest pressure and speed. PMID:27397738

  16. Evaluation of Pressure Stable Chip-to-Tube Fittings Enabling High-Speed Chip-HPLC with Mass Spectrometric Detection.

    PubMed

    Lotter, Carsten; Heiland, Josef J; Stein, Volkmar; Klimkait, Michael; Queisser, Marco; Belder, Detlev

    2016-08-01

    Appropriate chip-to-tube interfacing is an enabling technology for high-pressure and high-speed liquid chromatography on chip. For this purpose, various approaches, to connect pressure resistant glass chips with HPLC pumps working at pressures of up to 500 bar, were examined. Three side-port and one top-port connection approach were evaluated with regard to pressure stability and extra column band broadening. A clamp-based top-port approach enabled chip-HPLC-MS analysis of herbicides at the highest pressure and speed.

  17. Effects of chipping, grinding, and heat on survival of emerald ash borer, Agrilus planipennis (Coleoptera: Buprestidae), in chips.

    PubMed

    McCullough, Deborah G; Poland, Therese M; Cappaert, David; Clark, Erin L; Fraser, Ivich; Mastro, Victor; Smith, Sarah; Pell, Christopher

    2007-08-01

    The emerald ash borer, Agrilus planipennis Fairmaire (Coleoptera: Buprestidae), a phloem-feeding insect from Asia, was identified in 2002 as the cause of widespread ash (Fraxinus sp.) mortality in southeastern Michigan and Essex County, Ontario. Most larvae overwinter as nonfeeding prepupae in the outer sapwood or thick bark of large trees. In a series of studies, we evaluated effects of grinding, chipping, and heat treatment on survival of A. planipennis prepupae in ash material. Heavily infested ash bolts containing roughly 8,700 prepupae were processed by a horizontal grinder with either a 2.5- or 10-cm screen. There was no evidence of A. planipennis survival in chips processed with the 2.5-cm screen, but eight viable prepupae were recovered from chips processed with the 10-cm screen. We chiseled additional sentinel chips with prepupae from ash logs and buried 45 in each chip pile. In total, six prepupae in sentinel chips survived the winter, but we found no sign of adult A. planipennis emergence from the processed chips. Subsequently, we assessed prepupal survival in chips processed by a chipper or a horizontal grinder fit with 5-, 10-, or 12.7-cm screens. An estimated 1,565 A. planipennis prepupae were processed by each treatment. Chips from the chipper were shorter than chips from the grinder regardless of the screen size used. No live prepupae were found in chips produced by the chipper, but 21 viable prepupae were found in chips from the grinder. Infested wood and bark chips chiseled from logs were held in ovens at 25, 40, or 60 degrees C for 8, 24, or 48 h. Prepupal survival was consistently higher in wood chips than bark chips at 40 degrees C, whereas no prepupae survived exposure to 60 degrees C for eight or more hours. In a second study, prepupae in wood chips were exposed to 40, 45, 50, 55, or 60 degrees C for 20 or 120 min. Some prepupae survived 20 min of exposure to all temperatures. No prepupae survived exposure to 60 degrees C for 120 min, but 17

  18. Highly-integrated lab-on-chip system for point-of-care multiparameter analysis.

    PubMed

    Schumacher, Soeren; Nestler, Jörg; Otto, Thomas; Wegener, Michael; Ehrentreich-Förster, Eva; Michel, Dirk; Wunderlich, Kai; Palzer, Silke; Sohn, Kai; Weber, Achim; Burgard, Matthias; Grzesiak, Andrzej; Teichert, Andreas; Brandenburg, Albrecht; Koger, Birgit; Albers, Jörg; Nebling, Eric; Bier, Frank F

    2012-02-01

    A novel innovative approach towards a marketable lab-on-chip system for point-of-care in vitro diagnostics is reported. In a consortium of seven Fraunhofer Institutes a lab-on-chip system called "Fraunhofer ivD-platform" has been established which opens up the possibility for an on-site analysis at low costs. The system features a high degree of modularity and integration. Modularity allows the adaption of common and established assay types of various formats. Integration lets the system move from the laboratory to the point-of-need. By making use of the microarray format the lab-on-chip system also addresses new trends in biomedicine. Research topics such as personalized medicine or companion diagnostics show that multiparameter analyses are an added value for diagnostics, therapy as well as therapy control. These goals are addressed with a low-cost and self-contained cartridge, since reagents, microfluidic actuators and various sensors are integrated within the cartridge. In combination with a fully automated instrumentation (read-out and processing unit) a diagnostic assay can be performed in about 15 min. Via a user-friendly interface the read-out unit itself performs the assay protocol, data acquisition and data analysis. So far, example assays for nucleic acids (detection of different pathogens) and protein markers (such as CRP and PSA) have been established using an electrochemical read-out based on redoxcycling or an optical read-out based on total internal reflectance fluorescence (TIRF). It could be shown that the assay performance within the cartridge is similar to that found for the same assay in a microtiter plate. Furthermore, recent developments are the integration of sample preparation and polymerase chain reaction (PCR) on-chip. Hence, the instrument is capable of providing heating-and-cooling cycles necessary for DNA-amplification. In addition to scientific aspects also the production of such a lab-on-chip system was part of the development since

  19. Highly-integrated lab-on-chip system for point-of-care multiparameter analysis.

    PubMed

    Schumacher, Soeren; Nestler, Jörg; Otto, Thomas; Wegener, Michael; Ehrentreich-Förster, Eva; Michel, Dirk; Wunderlich, Kai; Palzer, Silke; Sohn, Kai; Weber, Achim; Burgard, Matthias; Grzesiak, Andrzej; Teichert, Andreas; Brandenburg, Albrecht; Koger, Birgit; Albers, Jörg; Nebling, Eric; Bier, Frank F

    2012-02-01

    A novel innovative approach towards a marketable lab-on-chip system for point-of-care in vitro diagnostics is reported. In a consortium of seven Fraunhofer Institutes a lab-on-chip system called "Fraunhofer ivD-platform" has been established which opens up the possibility for an on-site analysis at low costs. The system features a high degree of modularity and integration. Modularity allows the adaption of common and established assay types of various formats. Integration lets the system move from the laboratory to the point-of-need. By making use of the microarray format the lab-on-chip system also addresses new trends in biomedicine. Research topics such as personalized medicine or companion diagnostics show that multiparameter analyses are an added value for diagnostics, therapy as well as therapy control. These goals are addressed with a low-cost and self-contained cartridge, since reagents, microfluidic actuators and various sensors are integrated within the cartridge. In combination with a fully automated instrumentation (read-out and processing unit) a diagnostic assay can be performed in about 15 min. Via a user-friendly interface the read-out unit itself performs the assay protocol, data acquisition and data analysis. So far, example assays for nucleic acids (detection of different pathogens) and protein markers (such as CRP and PSA) have been established using an electrochemical read-out based on redoxcycling or an optical read-out based on total internal reflectance fluorescence (TIRF). It could be shown that the assay performance within the cartridge is similar to that found for the same assay in a microtiter plate. Furthermore, recent developments are the integration of sample preparation and polymerase chain reaction (PCR) on-chip. Hence, the instrument is capable of providing heating-and-cooling cycles necessary for DNA-amplification. In addition to scientific aspects also the production of such a lab-on-chip system was part of the development since

  20. Validation study for using lab-on-chip technology for Coxiella burnetii multi-locus-VNTR-analysis (MLVA) typing: application for studying genotypic diversity of strains from domestic ruminants in France.

    PubMed

    Prigent, Myriam; Rousset, Elodie; Yang, Elise; Thiéry, Richard; Sidi-Boumedine, Karim

    2015-01-01

    Coxiella burnetii, the etiologic bacterium of Q fever zoonosis, is still difficult to control. Ruminants are often carriers and involved in human epidemics. MLVA is a promising genotyping method for molecular epidemiology. Different techniques are used to resolve the MLVA band profiles such as electrophoresis on agarose gels, capillary electrophoresis or using the microfluidic Lab-on-Chip system. In this study, system based on microfluidics electrophoresis with Lab-on-Chip technology was assessed and applied on DNA field samples to investigate the genotypic diversity of C. burnetii strains circulating in France. The Lab-on-Chip technology was first compared to agarose gel electrophoresis. Subsequently, the set-up Lab-on-Chip technology was applied on 97 samples collected from ruminants in France using the 17 markers previously described. A discordance rate of 27% was observed between Lab-on-Chip and agarose gel electrophoresis. These discrepancies were checked and resolved by sequencing. The cluster analysis revealed classification based on host species and/or geographic origin criteria. Moreover, the circulation of different genotypic strains within the same farm was also observed. In this study, MLVA with Lab-on-Chip technology was shown to be more accurate, reproducible, user friendly and safer than gel electrophoresis. It also provides an extended data set from French ruminant C. burnetii circulating strains useful for epidemiological investigations. Finally, it raises some questions regarding the standardization and harmonization of C. burnetii MLVA genotyping.

  1. DNA Damage, DNA Repair, Aging, and Neurodegeneration.

    PubMed

    Maynard, Scott; Fang, Evandro Fei; Scheibye-Knudsen, Morten; Croteau, Deborah L; Bohr, Vilhelm A

    2015-09-18

    Aging in mammals is accompanied by a progressive atrophy of tissues and organs, and stochastic damage accumulation to the macromolecules DNA, RNA, proteins, and lipids. The sequence of the human genome represents our genetic blueprint, and accumulating evidence suggests that loss of genomic maintenance may causally contribute to aging. Distinct evidence for a role of imperfect DNA repair in aging is that several premature aging syndromes have underlying genetic DNA repair defects. Accumulation of DNA damage may be particularly prevalent in the central nervous system owing to the low DNA repair capacity in postmitotic brain tissue. It is generally believed that the cumulative effects of the deleterious changes that occur in aging, mostly after the reproductive phase, contribute to species-specific rates of aging. In addition to nuclear DNA damage contributions to aging, there is also abundant evidence for a causative link between mitochondrial DNA damage and the major phenotypes associated with aging. Understanding the mechanistic basis for the association of DNA damage and DNA repair with aging and age-related diseases, such as neurodegeneration, would give insight into contravening age-related diseases and promoting a healthy life span.

  2. A high-speed, high-performance on-chip integrated reverse transcription (RT)-microchip.

    PubMed

    Lee, Hwanyong; Han, Nari; Choi, In-Hak; Han, Ki-Ho

    2013-02-01

    This report introduces an on-chip integrated reverse transcription (RT)-microchip, which includes two genetic functionalities of RNA extraction and cDNA synthesis. In the RNA extraction compartment, RNA is extracted from peripheral blood lysate within 1 min, by lateral magnetophoresis using magnetic oligo-dT beads. The extracted RNA is then collected and used directly to produce cDNA in the cDNA synthesis microchamber, which is monolithically integrated with the RNA extraction compartment. To verify the superiority of the proposed RT-microchip, RT-PCR amplification was performed using cDNA harvested from the RT-microchip, and the results were compared with those obtained using typical RNA extraction methods such as a silica matrix column and magnetic oligo-dT beads. The RT-PCR amplification results using 100 μl of blood showed that the intensity of the bands in gel electrophoresis of the RT-microchip was 2-fold stronger than that of the silica matrix column and 2.65-fold stronger than that of the magnetic oligo-dT beads. The results demonstrate that the RT-microchip technique is the most sensitive of the tested methods.

  3. CHIP Is an Essential Determinant of Neuronal Mitochondrial Stress Signaling

    PubMed Central

    Palubinsky, Amy M.; Stankowski, Jeannette N.; Kale, Alixandra C.; Codreanu, Simona G.; Singer, Robert J.; Liebler, Daniel C.; Stanwood, Gregg D.

    2015-01-01

    Abstract Aims: Determine the mechanism by which C-terminus of HSC70-interacting protein (CHIP) induction alters neuronal survival under conditions of mitochondrial stress induced by oxygen glucose deprivation. Results: We report that animals deficient in the E3 ubiquitin ligase, CHIP, have high baseline levels of central nervous system protein oxidation and lipid peroxidation, reduced antioxidant defenses, and decreased energetic status. Stress-associated molecules typically linked to Parkinson's disease such as the mitochondrial kinase, PTEN-inducible putative kinase 1 (PINK1), and another E3 ligase, Parkin, are upregulated in brains from CHIP knockout (KO) animals. Utilizing a novel biotin–avidin capture technique, we found that the oxidation status of Parkin and the mitochondrial fission protein, dynamin-related protein 1 (Drp1), are altered in a CHIP-dependent manner. We also found that following oxygen–glucose deprivation (OGD), the expression of CHIP, PINK1, and the autophagic marker, LC3, increase and there is activation of the redox-sensitive kinase p66shc. Under conditions of OGD, CHIP relocalizes from the cytosol to mitochondria. Mitochondria from CHIP KO mice have profound impairments in stress response induced by calcium overload, resulting in accelerated permeability transition activity. While CHIP-deficient neurons are morphologically intact, they are more susceptible to OGD consistent with a previously unknown neuroprotective role for CHIP in maintaining mitochondrial homeostasis. Innovation: CHIP relocalization to the mitochondria is essential for the regulation of mitochondrial integrity and neuronal survival following OGD. Conclusions: CHIP is an essential regulator of neuronal bioenergetics and redox tone. Altering the expression of this protein has profound effects on neuronal survival when cells are exposed to OGD. Antioxid. Redox Signal. 23, 535–549. PMID:25602369

  4. DNA from plant mitochondria.

    PubMed

    Suyama, Y; Bonner, W D

    1966-03-01

    DNA WAS ISOLATED FROM A MITOCHONDRIAL FRACTION OF EACH OF THE FOLLOWING PLANT MATERIALS: Mung bean (Phaseolus aureus) etiolated hypocotyl; turnip (Brassica rapa) root; sweet potato (Ipomoea batatas) root; and onion (Allium cepa) bulb. It was found that all of these mitochondrial fractions contained DNA, the densities of which were identical (rho=1.706 g.cm(-3)). An additional DNA (rho=1.695) band found in the mitochondrial fraction of Brassica rapa, was identical to DNA separately isolated from the chloroplast-rich fraction. The origin of the second DNA from Allium mitochondrial fraction was not identified.Contrary to the identity of the mitochondrial DNA, DNA from nuclear fractions differed not only with each other but from the corresponding mitochondrial DNA.DNA from Phaseolus and Brassica mitochondria showed the hyperchromicity characteristic of double stranded, native DNA upon heating; Tm's in 0.0195 Na(+) were the same; 72.0 degrees . The amount of DNA within the mitochondrion of Phaseolus was estimated to be 5.0 x 10(-10) mug; this estimate was made by isolating the mitochondrial DNA concomitantly with the known amount of added (15)N(2)H B. subtilis DNA (rho=1.740). Approximately the same amount of DNA was present in the mitochondrion of Brassica or Ipomoea.

  5. Polymerase/DNA interactions and enzymatic activity: multi-parameter analysis with electro-switchable biosurfaces

    NASA Astrophysics Data System (ADS)

    Langer, Andreas; Schräml, Michael; Strasser, Ralf; Daub, Herwin; Myers, Thomas; Heindl, Dieter; Rant, Ulrich

    2015-07-01

    The engineering of high-performance enzymes for future sequencing and PCR technologies as well as the development of many anticancer drugs requires a detailed analysis of DNA/RNA synthesis processes. However, due to the complex molecular interplay involved, real-time methodologies have not been available to obtain comprehensive information on both binding parameters and enzymatic activities. Here we introduce a chip-based method to investigate polymerases and their interactions with nucleic acids, which employs an electrical actuation of DNA templates on microelectrodes. Two measurement modes track both the dynamics of the induced switching process and the DNA extension simultaneously to quantitate binding kinetics, dissociation constants and thermodynamic energies. The high sensitivity of the method reveals previously unidentified tight binding states for Taq and Pol I (KF) DNA polymerases. Furthermore, the incorporation of label-free nucleotides can be followed in real-time and changes in the DNA polymerase conformation (finger closing) during enzymatic activity are observable.

  6. Engineering molecularly-active nanoplasmonic surfaces for DNA detection via colorimetry and Raman scattering

    NASA Astrophysics Data System (ADS)

    Heydari, Esmaeil; Mabbott, Samuel; Thompson, David; Graham, Duncan; Cooper, Jonathan M.; Clark, Alasdair W.

    2016-03-01

    We report a novel nanophotonic biosensor surface capable of both colorimetric detection and Raman-scattered detection of DNA infection markers at extreme sensitivities. Combining direct-write lithography, dip-pen nanolithography based DNA patterning, and molecular self-assembly, we create molecularly-active plasmonic nanostructures onto which metallic nanoparticles are located via DNA-hybridization. Arraying these structures enables optical surfaces that change state when contacted by specific DNA sequences; shifting the surface color while simultaneously generating strong Raman-scattering signals. Patterning the DNA markers onto the plasmonic surface as micro-scale symbols results in easily identifiable color shifts, making this technique applicable to multiplexed lab-on-a-chip and point-of-care diagnostic applications.

  7. Polymerase/DNA interactions and enzymatic activity: multi-parameter analysis with electro-switchable biosurfaces

    PubMed Central

    Langer, Andreas; Schräml, Michael; Strasser, Ralf; Daub, Herwin; Myers, Thomas; Heindl, Dieter; Rant, Ulrich

    2015-01-01

    The engineering of high-performance enzymes for future sequencing and PCR technologies as well as the development of many anticancer drugs requires a detailed analysis of DNA/RNA synthesis processes. However, due to the complex molecular interplay involved, real-time methodologies have not been available to obtain comprehensive information on both binding parameters and enzymatic activities. Here we introduce a chip-based method to investigate polymerases and their interactions with nucleic acids, which employs an electrical actuation of DNA templates on microelectrodes. Two measurement modes track both the dynamics of the induced switching process and the DNA extension simultaneously to quantitate binding kinetics, dissociation constants and thermodynamic energies. The high sensitivity of the method reveals previously unidentified tight binding states for Taq and Pol I (KF) DNA polymerases. Furthermore, the incorporation of label-free nucleotides can be followed in real-time and changes in the DNA polymerase conformation (finger closing) during enzymatic activity are observable. PMID:26174478

  8. Prostaglandin E₂ increases fibroblast gene-specific and global DNA methylation via increased DNA methyltransferase expression.

    PubMed

    Huang, Steven K; Scruggs, Anne M; Donaghy, Jake; McEachin, Richard C; Fisher, Aaron S; Richardson, Bruce C; Peters-Golden, Marc

    2012-09-01

    Although alterations in DNA methylation patterns have been associated with specific diseases and environmental exposures, the mediators and signaling pathways that direct these changes remain understudied. The bioactive lipid mediator prostaglandin E(2) (PGE(2)) has been shown to exert a myriad of effects on cell survival, proliferation, and differentiation. Here, we report that PGE(2) also signals to increase global DNA methylation and DNA methylation machinery in fibroblasts. HumanMethylation27 BeadChip array analysis of primary fetal (IMR-90) and adult lung fibroblasts identified multiple genes that were hypermethylated in response to PGE(2). PGE(2), compared with nontreated controls, increased expression and activity (EC(50)∼10(7) M) of one specific isoform of DNA methyltransferase, DNMT3a. Silencing of DNMT3a negated the ability of PGE(2) to increase DNMT activity. The increase in DNMT3a expression was mediated by PGE(2) signaling via its E prostanoid 2 receptor and the second messenger cAMP. PGE(2), compared with the untreated control, increased the expression and activity of Sp1 and Sp3 (EC(50)∼3×10(7) M), transcription factors known to increase DNMT3a expression, and inhibition of these transcription factors abrogated the PGE(2) increase of DNMT3a expression. These findings were specific to fibroblasts, as PGE(2) decreased DNMT1 and DNMT3a expression in RAW macrophages. Taken together, these findings establish that DNA methylation is regulated by a ubiquitous bioactive endogenous mediator. Given that PGE(2) biosynthesis is modulated by environmental toxins, various disease states, and commonly used pharmacological agents, these findings uncover a novel mechanism by which alterations in DNA methylation patterns may arise in association with disease and certain environmental exposures.

  9. Rapid detection of aflatoxigenic Aspergillus sp. in herbal specimens by a simple, bendable, paper-based lab-on-a-chip.

    PubMed

    Chaumpluk, Piyasak; Plubcharoensook, Pattra; Prasongsuk, Sehanat

    2016-06-01

    Postharvest herbal product contamination with mycotoxins and mycotoxin-producing fungi represents a potentially carcinogenic hazard. Aspergillus flavus is a major cause of this issue. Available mold detection methods are PCR-based and rely heavily on laboratories; thus, they are unsuitable for on-site monitoring. In this study, a bendable, paper-based lab-on-a-chip platform was developed to rapidly detect toxigenic Aspergillus spp. DNA. The 3.0-4.0 cm(2) chip is fabricated using Whatman™ filter paper, fishing line and a simple plastic lamination process and has nucleic acid amplification and signal detection components. The Aspergillus assay specifically amplifies the aflatoxin biosynthesis gene, aflR, using loop-mediated isothermal amplification (LAMP); hybridization between target DNA and probes on blue silvernanoplates (AgNPls) yields colorimetric results. Positive results are indicated by the detection pad appearing blue due to dispersed blue AgNPls; negative results are indicated by the detection pad appearing colorless or pale yellow due to probe/target DNA hybridization and AgNPls aggregation. Assay completion requires less than 40 min, has a limit of detection (LOD) of 100 aflR copies, and has high specificity (94.47%)and sensitivity (100%). Contamination was identified in 14 of 32 herbal samples tested (43.75%). This work demonstrates the fabrication of a simple, low-cost, paper-based lab-on-a-chip platform suitable for rapid-detection applications. PMID:27168276

  10. High-Throughput Screening Platform for Engineered Nanoparticle-Mediated Genotoxicity Using CometChip Technology

    PubMed Central

    2015-01-01

    The likelihood of intentional and unintentional engineered nanoparticle (ENP) exposure has dramatically increased due to the use of nanoenabled products. Indeed, ENPs have been incorporated in many useful products and have enhanced our way of life. However, there are many unanswered questions about the consequences of nanoparticle exposures, in particular, with regard to their potential to damage the genome and thus potentially promote cancer. In this study, we present a high-throughput screening assay based upon the recently developed CometChip technology, which enables evaluation of single-stranded DNA breaks, abasic sites, and alkali-sensitive sites in cells exposed to ENPs. The strategic microfabricated, 96-well design and automated processing improves efficiency, reduces processing time, and suppresses user bias in comparison to the standard comet assay. We evaluated the versatility of this assay by screening five industrially relevant ENP exposures (SiO2, ZnO, Fe2O3, Ag, and CeO2) on both suspension human lymphoblastoid (TK6) and adherent Chinese hamster ovary (H9T3) cell lines. MTT and CyQuant NF assays were employed to assess cellular viability and proliferation after ENP exposure. Exposure to ENPs at a dose range of 5, 10, and 20 μg/mL induced dose-dependent increases in DNA damage and cytotoxicity. Genotoxicity profiles of ZnO > Ag > Fe2O3 > CeO2 > SiO2 in TK6 cells at 4 h and Ag > Fe2O3 > ZnO > CeO2 > SiO2 in H9T3 cells at 24 h were observed. The presented CometChip platform enabled efficient and reliable measurement of ENP-mediated DNA damage, therefore demonstrating the efficacy of this powerful tool in nanogenotoxicity studies. PMID:24617523

  11. Novel genomic island modifies DNA with 7-deazaguanine derivatives

    PubMed Central

    Thiaville, Jennifer J.; Kellner, Stefanie M.; Yuan, Yifeng; Hutinet, Geoffrey; Thiaville, Patrick C.; Jumpathong, Watthanachai; Mohapatra, Susovan; Brochier-Armanet, Celine; Letarov, Andrey V.; Hillebrand, Roman; Malik, Chanchal K.; Rizzo, Carmelo J.; Dedon, Peter C.; de Crécy-Lagard, Valérie

    2016-01-01

    The discovery of ∼20-kb gene clusters containing a family of paralogs of tRNA guanosine transglycosylase genes, called tgtA5, alongside 7-cyano-7-deazaguanine (preQ0) synthesis and DNA metabolism genes, led to the hypothesis that 7-deazaguanine derivatives are inserted in DNA. This was established by detecting 2’-deoxy-preQ0 and 2’-deoxy-7-amido-7-deazaguanosine in enzymatic hydrolysates of DNA extracted from the pathogenic, Gram-negative bacteria Salmonella enterica serovar Montevideo. These modifications were absent in the closely related S. enterica serovar Typhimurium LT2 and from a mutant of S. Montevideo, each lacking the gene cluster. This led us to rename the genes of the S. Montevideo cluster as dpdA-K for 7-deazapurine in DNA. Similar gene clusters were analyzed in ∼150 phylogenetically diverse bacteria, and the modifications were detected in DNA from other organisms containing these clusters, including Kineococcus radiotolerans, Comamonas testosteroni, and Sphingopyxis alaskensis. Comparative genomic analysis shows that, in Enterobacteriaceae, the cluster is a genomic island integrated at the leuX locus, and the phylogenetic analysis of the TgtA5 family is consistent with widespread horizontal gene transfer. Comparison of transformation efficiencies of modified or unmodified plasmids into isogenic S. Montevideo strains containing or lacking the cluster strongly suggests a restriction–modification role for the cluster in Enterobacteriaceae. Another preQ0 derivative, 2’-deoxy-7-formamidino-7-deazaguanosine, was found in the Escherichia coli bacteriophage 9g, as predicted from the presence of homologs of genes involved in the synthesis of the archaeosine tRNA modification. These results illustrate a deep and unexpected evolutionary connection between DNA and tRNA metabolism. PMID:26929322

  12. LCAT DNA shearing.

    PubMed

    Okabe, Yuka; Lee, Abraham P

    2014-04-01

    We present a novel method to fragment DNA by using lateral cavity acoustic transducers (LCATs). DNA solution is placed within a microfluidic device containing LCATs. The LCATs cause microstreaming, which fragments DNA within the solution without any need for purification or downstream processing. The LCAT-based DNA fragmentation method offers an easy-to-use, low-cost, low-energy way to fragment DNA that is amenable to integration on microfluidic platforms to further automate DNA processing. Furthermore, the LCAT microdevice requires less than 10 µL of sample, and no external equipment is needed besides a piezoelectric transducer. PMID:23850863

  13. Real-time forensic DNA analysis at a crime scene using a portable microchip analyzer.

    PubMed

    Liu, Peng; Yeung, Stephanie H I; Crenshaw, Karin A; Crouse, Cecelia A; Scherer, James R; Mathies, Richard A

    2008-09-01

    An integrated lab-on-a-chip system has been developed and successfully utilized for real-time forensic short tandem repeat (STR) analysis. The microdevice comprises a 160-nL polymerase chain reaction reactor with an on-chip heater and a temperature sensor for thermal cycling, microvalves for fluidic manipulation, a co-injector for sizing standard injection, and a 7-cm-long separation channel for capillary electrophoretic analysis. A 9-plex autosomal STR typing system consisting of amelogenin and eight combined DNA index system (CODIS) core STR loci has been constructed and optimized for this real-time human identification study. Reproducible STR profiles of control DNA samples are obtained in 2h and 30min with DNA required for a complete DNA profile is 100 copies. To critically evaluate the capabilities of our portable microsystem as well as its compatibility with crime scene investigation processes, real-time STR analyses were carried out at a mock crime scene prepared by the Palm Beach County Sheriff's Office (PBSO). Blood stain sample collection, DNA extraction, and STR analyses on the portable microsystem were conducted in the field, and a successful "mock" CODIS hit was generated on the suspect's sample within 6h. This demonstration of on-site STR analysis establishes the feasibility of real-time DNA typing to identify the contributor of probative biological evidence at a crime scene and for real-time human identification.

  14. DNA methylome profiling of maternal peripheral blood and placentas reveal potential fetal DNA markers for non-invasive prenatal testing.

    PubMed

    Xiang, Yuqian; Zhang, Junyu; Li, Qiaoli; Zhou, Xinyao; Wang, Teng; Xu, Mingqing; Xia, Shihui; Xing, Qinghe; Wang, Lei; He, Lin; Zhao, Xinzhi

    2014-09-01

    Utilizing epigenetic (DNA methylation) differences to differentiate between maternal peripheral blood (PBL) and fetal (placental) DNA has been a promising strategy for non-invasive prenatal testing (NIPT). However, the differentially methylated regions (DMRs) have yet to be fully ascertained. In the present study, we performed genome-wide comparative methylome analysis between maternal PBL and placental DNA from pregnancies of first trimester by methylated DNA immunoprecipitation-sequencing (MeDIP-Seq) and Infinium HumanMethylation450 BeadChip assays. A total of 36 931 DMRs and 45 804 differentially methylated sites (DMSs) covering the whole genome, exclusive of the Y chromosome, were identified via MeDIP-Seq and Infinium 450k array, respectively, of which 3759 sites in 2188 regions were confirmed by both methods. Not only did we find the previously reported potential fetal DNA markers in our identified DMRs/DMSs but also we verified fully the identified DMRs/DMSs in the validation round by MassARRAY EpiTYPER. The screened potential fetal DNA markers may be used for NIPT on aneuploidies and other chromosomal diseases, such as cri du chat syndrome and velo-cardio-facial syndrome. In addition, these potential markers may have application in the early diagnosis of placental dysfunction, such as pre-eclampsia. PMID:24996894

  15. A method for evaluation of the quality of DNA microarray spots.

    PubMed

    Boa, Zhang; Ma, Wen-Li; Hu, Zi-You; Rong, Shi; Shi, Yan-Bin; Zheng, Wen-Ling

    2002-09-30

    To establish a method to evaluate the quality of the printed microarray and DNA fragments' immobilization. The target gene fragments that were made with the restriction display PCR (RD-PCR) technique were printed on a superamine modified glass slide, then immobilized with UV cross-linking and heat. This chip was hybridized with universal primers that were labeled with cy3-dUTP, as well as cDNA that was labeled with cy3-dCTP, as the conventional protocol. Most of the target gene fragments on the chip showed positive signals, but the negative control showed no signal, and vice versa. We established a method that enables an effective evaluation of the quality of the microarrays.

  16. Selection of functional human sperm with higher DNA integrity and fewer reactive oxygen species

    PubMed Central

    Asghar, Waseem; Velasco, Vanessa; Kingsley, James L.; Shoukat, Muhammad S.; Shafiee, Hadi; Anchan, Raymond M.; Mutter, George L.; Tüzel, Erkan; Demirci, Utkan

    2014-01-01

    Fertilization and reproduction are central to the survival and propagation of a species. Couples who cannot reproduce naturally have to undergo in vitro clinical procedures. An integral part of these clinical procedures includes isolation of healthy sperm from raw semen. Existing sperm sorting methods are not efficient and isolate sperm having high DNA fragmentation and reactive oxygen species, and suffer from multiple manual steps and variations between embryologists. Inspired by in vivo natural sperm sorting mechanisms where vaginal mucus becomes less viscous to form microchannels to guide sperm towards egg, we present a chip that efficiently sorts healthy, motile and morphologically normal sperm without centrifugation. Higher percentage of sorted sperm show significantly lesser reactive oxygen species and DNA fragmentation than the conventional swim-up method. The presented chip is an easy-to-use high throughput sperm sorter that provides standardized sperm sorting assay with less reliance on embryologist’s skills, facilitating reliable operational steps. PMID:24753434

  17. Wireless spread-spectrum telesensor chip with synchronous digital architecture

    DOEpatents

    Smith, Stephen F.; Turner, Gary W.; Wintenberg, Alan L.; Emery, Michael Steven

    2005-03-08

    A fully integrated wireless spread-spectrum sensor incorporating all elements of an "intelligent" sensor on a single circuit chip is capable of telemetering data to a receiver. Synchronous control of all elements of the chip provides low-cost, low-noise, and highly robust data transmission, in turn enabling the use of low-cost monolithic receivers.

  18. The three-dimensional structure of human erythrocyte aquaporin CHIP.

    PubMed

    Walz, T; Smith, B L; Agre, P; Engel, A

    1994-07-01

    Water-permeable membranes of several plant and mammalian tissues contain specific water channel proteins, the 'aquaporins'. The best characterized aquaporin is CHIP, a 28 kDa red blood cell channel-forming integral protein. Isolated CHIP and Escherichia coli lipids may be assembled into 2-D crystals for structural analyses. Here we present (i) a structural characterization of the solubilized CHIP oligomers, (ii) projections of CHIP arrays after negative staining or metal-shadowing, and (iii) the 3-D structure at 1.6 nm resolution. Negatively stained CHIP oligomers exhibited a side length of 6.9 nm with four-fold symmetry, and a mass of 202 +/- 3 kDa determined by scanning transmission electron microscopy. Reconstituted into lipid bilayers, CHIP formed 2-D square lattices with unit cell dimensions a = b = 9.6 nm and a p422(1) symmetry. The 3-D map revealed that CHIP tetramers contain central stain-filled depressions about the fourfold axis. These cavities extend from both sides into the transbilayer domain of the molecule leaving only a thin barrier to be penetrated by the water pores. Although CHIP monomers behave as independent pores, we propose that their particular structure requires tetramerization for stable integration into the bilayer. PMID:7518771

  19. 'Intelligent Memory Chips' Give Fully Programmable Synaptic Weights

    NASA Astrophysics Data System (ADS)

    Morton, Steven G.

    1989-09-01

    A fundamental stumbling block - defining a new set of extremely powerful and flexible building blocks with which to build neurocomputers - has recently been removed by Oxford Computer. The result is a family of digital, memory-plus-processor chips, or "Intelligent Memory Chips". These chips combine a high-capacity memory with massively parallel, slice-type processor logic. Unlike common memory chips that only store information, Intelligent Memory Chips perform intensive computations upon matrices they store. As a result, neural networks with fully programmable, signed synaptic weights can be built. The weights are modified as easily, precisely and stably as writing data into ordinary memory chips. Many forms of matrix-vector multipliers, 1- and 2-dimensional convolvers, and Fast Fourier and other transformers can be built as well to implement classical digital signal processing, pattern recognition, adaptive control and 3-dimensional graphics structures. Multiple Intelligent Memory Chips work together to provide the precision, matrix size and performance desired. Extremely large numbers of densely interconnected, artificial neurons in many layers can be provided. Networks easily interface to existing, non-neural machines. Network performance ranging from tens-of-billions to tens-of-trillions of operations per second may be built using current to near term semiconductor technology. Initial chips are being built using 1-micron, silicon CMOS and static RAM technology. The impacts of alternative memory technologies, and improvements in memory and fabrication technology are also discussed.

  20. An addressable cell array for a platform of biosensor chips

    NASA Astrophysics Data System (ADS)

    Yang, Seungkyoung; Choi, Soo-hee; Jung, Moon Youn; Song, Kibong; Park, Jeong Won

    2013-05-01

    In order to detect interested matters in fields, various lab-on-a-chips where chemical, physical, or biological sensors are loaded have been developed. eNOSE can be a representative example among them. Because animals can sense 300~1000 different chemicals by olfactory system - smell -, the olfactory system has been spotlighted as new materials in the field of sensing. Those investigations, however, are usually focused on how to detect signals from the olfactory neurons or receptors loaded on chips and enhance sensing efficacy of chips. Therefore, almost of those chips are designed for only one material sensing. Multi-sensing using multi-channels will be needed when the olfactory systems are adopted well on chips. For multiple sensing, we developed an addressable cell array. The chip has 38 cell-chambers arranged in a circle shape and different cell types of thirty eight can be allocated with specific addresses on the chip without any complex valve system. In order to confirm the cell addressing, we loaded EGFP-transfected and empty vector-transfected HEK293a cells into inlets of the cell array in a planned address and those cells were positioned into each chamber by brief aspiration. The arrayed cells were confirmed as a specific pattern through EGFP and nuclei staining. This cell array which can generate address of sensor materials like cells with their own specification is expected to be applied to a platform for a biosensor chip at various sensing fields.