Дисертації з теми "Digital polymerase chain reaction"
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Al-hashimi, Sora. "Kvantifiering med digital droplet polymerase chain reaction av gyrA-genen med och utan mutationen S83L." Thesis, Örebro universitet, Institutionen för hälsovetenskaper, 2021. http://urn.kb.se/resolve?urn=urn:nbn:se:oru:diva-92989.
Повний текст джерелаThe most common form of cancer in men is prostate cancer, with 10,000 new deaths annually. In prostate cancer diagnosis, prostate biopsy is performed. To reduce the risk of complications in connection with biopsy, a single dose of the antibiotic drug Ciprofloxacin is given in Sweden. The proportion of bacteria that are resistant to ciprofloxacin has increased. For the detection of gene mutations that cause antibiotic resistance, droplet digital PCR (ddPCR) can be used. It is a method that provides an absolute quantification of a DNA sequence in a sample. It is based on water oil emulsion drop system. The purpose of this study was to optimize and validate a digital droplet PCR to detect and quantify the S83L mutation in the gyrA gene from faecal samples and to compare digital droplet results from study samples with culture results from resistance determination and the ration between the S83L allele and the wild-type allele in samples taken before and after biopsy. To validate the method, samples taken before and after biopsy were used from nine patients who had undergone a transrectal prostate biopsy and received a dose of ciprofloxacin or trimethoprim in connection with the procedure. The optimal annealing temperature was determined to be 60 °C and the optimal primer and probe concentrations were determined to be 1.2 µM and 0.4µM, respectively. These concentrations gave the lowest number of false positive droplets. The minimum detection level for S83L gyrA (EC40) was 160 copies/ml and for wild-type gyrA (EC108) it was 78 copies/ml. The results showed that both wild-type gyrA and S83L gyrA could be detected and quantified in rectal samples from all nine patients.
Poleti, Marcelo Lupion. "Análise morfométrica, radiográfica e molecular do processo de reparo alveolar após a terapia fotodinâmica antimicrobiana em ratos Wistar." Universidade de São Paulo, 2009. http://www.teses.usp.br/teses/disponiveis/25/25132/tde-03072009-105031/.
Повний текст джерелаIntroduction: Antibiotics resistance made the development of antimicrobial alternative techniques necessary. Consequently, others alternatives such as antimicrobial photodynamic therapy (PDT) have been studied. Objective: The objectives were to perform morphometric, radiographic and molecular analyses of alveolar repair process in rats after antimicrobial photodynamic therapy. Methods: Eighty-five rats were used in this study, divided according to the following groups: C: untreated socket; S+L: socket treatment with physiologic saline solution and low intensity laser therapy (660nm - 50J/cm2); ATO: socket treatment with topic application of toluidine blue-O (100 µg/mL) and ATO+L: socket treatment with topic application of toluidine blue-O (100µg/ml) and low intensity laser therapy (660nm - 50J/cm2). The animals were sacrificed at a postoperative period of 6, 15 and 28 days. Thermal variation was carried out in an animal to confirm the absence of Laser thermal effect on irradiation area. Quantitative and qualitative microscopic analyses were performed to evaluate the connective tissue, bone tissue, blood clot, blood vessel, inflammatory infiltrate and empty space. Fractal and Pixel radiographic analysis were performed. A quantitative analysis was performed using a RealTimePCR to evaluate the genes expression involved Collagen Type I (COL-I), vascular endothelial growth factor (VEGF), runt-related transcription factor 2 (RUNX2), osteocalcin (OCN), alkaline phosphatase (ALP), in the alveolar repair process. Results and Conclusions: Based in the results, it can be concluded that antimicrobial photodynamic therapy did not act negatively in the socket bone repair evaluation. Pixel values analysis could be used as indicators to evaluate the dry socket bone neoformation. The VEGF molecular marker could be used as angiogenesis indicator to evaluate the dry socket bone neoformation as well as alkaline phosphatase as a molecular marker in the early stage of bone neoformation.
Feugeas, Olivier. "Pcr (polymerase chain reaction) et vih." Lille 2, 1990. http://www.theses.fr/1990LIL2M264.
Повний текст джерелаVerhaegen, Monique Elise. "Novel approaches in quantitative polymerase chain reaction." Thesis, National Library of Canada = Bibliothèque nationale du Canada, 1998. http://www.collectionscanada.ca/obj/s4/f2/dsk1/tape10/PQDD_0021/MQ52489.pdf.
Повний текст джерелаChiou, Jeffrey Tsungshuan. "A novel capillary polymerase chain reaction machine." Thesis, Massachusetts Institute of Technology, 2001. http://hdl.handle.net/1721.1/8864.
Повний текст джерелаIncludes bibliographical references (p. 254-268).
I built a novel prototype capillary polymerase chain reaction machine. The purpose was to perform a single reaction as fast as possible with a reaction volume - 100 nl. The PCR mix is in the form of a 1 /1 droplet that moves between three heat zones inside of a 1 mm I.D. capillary filled with mineral oil via pneumatic actuation. A laser beam waveguides down the capillary until it strikes the drop, at which point it scatters. The scatter is picked up by a series of photodiodes to provide position feedback. Due to the efficient heat transfer arrangement, the drop can transition between different temperature steps in -2 seconds, which includes both drop motion and temperature equilibration. It was extensively tested in both 10-cycle and 30-cycle PCR, including nearly 200 successful 30-cycle runs. The 30-cycle PCR was typically 74% (as high as 78%) efficient, and took only 23 minutes. This compares well with existing machines in the literature.
by Jeffrey Tsungshuan Chiou.
Ph.D.
Clackson, Timothy Piers. "Antibody engineering using the polymerase chain reaction." Thesis, University of Cambridge, 1991. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.316695.
Повний текст джерелаLantz, Pär-G. "PCR-based detection of microorganisms in complex biological samples." Lund : Dept. of Applied Microbiology, Lund University, 1998. http://catalog.hathitrust.org/api/volumes/oclc/39178906.html.
Повний текст джерелаLinley, M. "The detection of polymerase inhibiting lesions using the polymerase arrest polymerase chain reaction assay." Thesis, Swansea University, 2004. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.637924.
Повний текст джерелаNebbali, M. "Human gene mapping using the polymerase chain reaction." Thesis, University of Nottingham, 1992. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.317395.
Повний текст джерелаBorges, Pinto Lais Izabel. "Alu-polymerase chain reaction genomic fingerprinting in neuroblastoma." Thesis, University of Newcastle Upon Tyne, 2001. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.366679.
Повний текст джерелаErill, Sagalés Ivan. "High-speed Polymerase chain reaction in CMOS-compatible chips." Doctoral thesis, Universitat Autònoma de Barcelona, 2002. http://hdl.handle.net/10803/3031.
Повний текст джерелаEn el transcurso de esta tesis doctoral, se ha llevado a cabo el desarrollo un proceso tecnológico común para la fabricación de DNA-chips multifunción (i.e. sistemas versátiles basados en PCR y electroforesis), poniendo un especial énfasis en la compatibilidad con los procesos CMOS estándar, a fin de conseguir desarrollar prototipos proto-industriales. Como demostrador de esta puesta a punto tecnológica, se han diseñado, fabricado y testado chips de PCR, y la PCR en chips ha sido optimizada con respecto a materiales de fabricación, metodologías de inserción/extracción, composición bioquímica de la mix de PCR, diferentes configuraciones de calentadores/sensores (Peltier/termopares vs. resistencias integradas) y la cinética de la reacción.
In the last decade of the twentieth century, the fields of µ-TAS and, more specifically, DNA-chips have acquired increasing importance in the microsystems arena. The main reason for this surge of interest lies in the advantages these new devices seek to bring forth: faster, cheaper and completely automated analyses, and also in the outbreak of novel analytical techniques (e.g. hybridization chips). In the particular case of DNA-chips, functional prototypes have been demonstrated for PCR, LCR, gel electrophoresis, di-electrophoresis, hybridization and various combinations of these techniques, whilst hybridization chips (mainly arrayer chips) have become a successful market application. But, even though a considerable amount of work has been carried out in these few years, much research is still required to address fundamental problems of DNA-chips.
In this doctoral work, a common-ground technological setup for the production of multifunction DNA-chips (i.e. PCR plus electrophoresis systems) has been laid down, placing strong emphasis in its compatibility with standard CMOS processes in order to produce proto-industrial prototypes. As a demonstrator of this technological setup, PCR-chips have been designed, manufactured and tested, and the chip PCR reaction has been optimized with respect to surface materials, insertion and extraction methods, biochemical mix composition, heater/sensor setups (Peltier/thermocouple vs. thin-film driven systems) and reaction kinetics.
Aydin, Gamze. "Detection Of Genetically Modified Maize Via Polymerase Chain Reaction." Master's thesis, METU, 2004. http://etd.lib.metu.edu.tr/upload/3/12605495/index.pdf.
Повний текст джерелаBallagi-Pordány, András. "Application of polymerase chain reaction (PCR) in veterinary virology /." Uppsala : Sveriges lantbruksuniv, 1995. http://epsilon.slu.se/avh/1995/91-576-4997-9.gif.
Повний текст джерелаWoolford, Alison Jane. "Use of polymerase chain reaction for detection of mycobacteria." Thesis, University of Surrey, 1995. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.308497.
Повний текст джерелаGurram, Neil (Neil K. ). "A mathematical model of polymerase chain reaction induced stutter." Thesis, Massachusetts Institute of Technology, 2016. http://hdl.handle.net/1721.1/106012.
Повний текст джерелаThis electronic version was submitted by the student author. The certified thesis is available in the Institute Archives and Special Collections.
Cataloged from student-submitted PDF version of thesis.
Includes bibliographical references (page 48).
This is a thesis on understanding stutter present in capillary electropherogram readouts as this methodology forms the basis of current DNA fingerprinting. The readouts come from taking samples of various initial template masses of DNA from different individuals, applying polymerase chain reaction (PCR) to the samples, and then running the amplified copies through capillary electrophoresis to produce a readout of peak heights corresponding to alleles on various loci. The alleles correspond to the number of repeats of microsatellites that are usually two to six base pairs in length called short tandem repeats (STRs); the number of repeats of various STRs defines a person's DNA fingerprint. This process introduces artifacts in measurement. Of particular interest in this thesis is stutter, the phenomenon where amplicons with fewer or greater number of STR repeats than the true allele count are generated as an artifact of the PCR. It is of interest to understand the source and nature for this stutter distribution for small starting masses, as it has ramifications on the ability to accurately determine a match between a DNA sample and a crime scene sample. Understanding the stutter distribution in this thesis is achieved through data analysis, probabilistic modeling, and statistics. We find that a mathematical model that combines stochastic effects from PCR with fluorescent noise explains the most significant features of the observed phenomena.
by Neil Gurram.
M. Eng.
Lewis, Monte. "Thermal cycling design alternatives for the polymerase chain reaction." College Park, Md. : University of Maryland, 2005. http://hdl.handle.net/1903/3061.
Повний текст джерелаThesis research directed by: Dept. of Mechanical Engineering. Title from t.p. of PDF. Includes bibliographical references. Published by UMI Dissertation Services, Ann Arbor, Mich. Also available in paper.
Way, Jaw-Shiow Chu. "Specific detection of Salmonella by multiplex polymerase chain reaction." Diss., The University of Arizona, 1993. http://hdl.handle.net/10150/186207.
Повний текст джерелаMuwonge, Abubaker. "Detection Of Genetically Modified Potatoes By The Polymerase Chain Reaction." Master's thesis, METU, 2005. http://etd.lib.metu.edu.tr/upload/12605783/index.pdf.
Повний текст джерелаWhite, Adam. "Development and application of microfluidic single-cell polymerase chain reaction." Thesis, University of British Columbia, 2015. http://hdl.handle.net/2429/55734.
Повний текст джерелаScience, Faculty of
Graduate
Tsang, Tsui-ying Stella, and 曾璀瑩. "Application of quantitative polymerase chain reaction in the diagnosisof thalassaemia." Thesis, The University of Hong Kong (Pokfulam, Hong Kong), 2006. http://hub.hku.hk/bib/B45010948.
Повний текст джерелаGamma, Torres Rafael Enrique. "Application of polymerase chain reaction to rhinovirus detection and analysis." Thesis, University of Essex, 1992. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.293589.
Повний текст джерелаKoc, Yasemin. "Optimization of continuous flow polymerase chain reaction with microfluidic reactors." Thesis, Imperial College London, 2008. http://hdl.handle.net/10044/1/8184.
Повний текст джерелаAllmann, Michael. "Applications of the polymerase chain reaction (PCR) in food chemistry /." [S.l.] : [s.n.], 1994. http://www.ub.unibe.ch/content/bibliotheken_sammlungen/sondersammlungen/dissen_bestellformular/index_ger.html.
Повний текст джерелаMello, Richard. "IDENTIFICATION OF DQ ALPHA POLYMORPHISM USING THE POLYMERASE CHAIN REACTION." VCU Scholars Compass, 1991. https://scholarscompass.vcu.edu/etd/5213.
Повний текст джерелаSonmezalp, C. Zeynep. "Detection Of Genetically Modified Insect Resistant Tomato Via Polymerase Chain Reaction." Master's thesis, METU, 2004. http://etd.lib.metu.edu.tr/upload/3/12605493/index.pdf.
Повний текст джерелаDuman, Zeynep. "Polymerase Chain Reaction (pcr) For Detection Of Borrelia Burgdorferi Sensu Lato." Master's thesis, METU, 2008. http://etd.lib.metu.edu.tr/upload/2/12609199/index.pdf.
Повний текст джерелаTsang, Tsui-ying Stella. "Application of quantitative polymerase chain reaction in the diagnosis of thalassaemia /." View the Table of Contents & Abstract, 2006. http://sunzi.lib.hku.hk/hkuto/record/B36433895.
Повний текст джерелаRos, Bascuñana Carlos. "Diagnostic application of the polymerase chain reaction (PCR) in veterinary microbiology /." Uppsala : Swedish Univ. of Agricultural Sciences (Sveriges lantbruksuniv.), 1997. http://epsilon.slu.se/avh/1997/91-576-5247-3.gif.
Повний текст джерелаWalker, Ken R. "Rapid detection of Listeria monocytogenes in salad by polymerase chain reaction." Auburn, Ala, 2005. http://repo.lib.auburn.edu/2005%20Summer/master's/WALKER_KEN_28.pdf.
Повний текст джерелаPhaneuf, Christopher. "Infrared laser-mediated polymerase chain reaction in a polymer microfluidic device." Diss., Georgia Institute of Technology, 2014. http://hdl.handle.net/1853/53068.
Повний текст джерелаTully, Gillian. "DNA profiling for forensic identification : evaluation of polymerase chain reaction methods." Thesis, Cardiff University, 1996. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.264882.
Повний текст джерелаDiss, Timothy Charles. "The polymerase chain reaction in the characterisation and diagnosis of lymphomas." Thesis, University College London (University of London), 1996. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.338653.
Повний текст джерелаOrganji, Sameer R. A. "Detection of #Beta#-lactam resistant Streptococcus pneumoniae by polymerase chain reaction." Thesis, University of Bristol, 1998. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.297824.
Повний текст джерелаSim, Steven Poh Chuen. "An integrated chip-based device for droplet-flow polymerase chain reaction." Thesis, Imperial College London, 2016. http://hdl.handle.net/10044/1/56111.
Повний текст джерелаLiao, Yu-Hua. "Polymerase chain reaction based cloning of acetate kinase in propionibacterium acidipropionici." The Ohio State University, 2004. http://rave.ohiolink.edu/etdc/view?acc_num=osu1072778140.
Повний текст джерелаHolladay, Ervin Blair. "Viral gene detection in oral neoplasms using the polymerase chain reaction." The Ohio State University, 1991. http://rave.ohiolink.edu/etdc/view?acc_num=osu1345129906.
Повний текст джерелаHolladay, E. Blair. "Viral gene detection in oral neoplasms using the polymerase chain reaction /." The Ohio State University, 1991. http://rave.ohiolink.edu/etdc/view?acc_num=osu1487694702781994.
Повний текст джерелаLiao, Yu-Hua. "Polymerase chain reaction based cloning of acetate kinase in Propionibacterium acidipropionici /." Connect to this title online, 2003. http://rave.ohiolink.edu/etdc/view?acc%5Fnum=osu1072778140.
Повний текст джерелаLiu, Tingting. "Electrokinetic Real-Time Polymerase Chain Reaction Toward Point-Of-Care Diagnosis." Diss., The University of Arizona, 2015. http://hdl.handle.net/10150/579083.
Повний текст джерелаLiu, Pang-I., and 劉邦奕. "Design and Fabrication of Microfluidic Devices for Digital Polymerase Chain Reaction." Thesis, 2018. http://ndltd.ncl.edu.tw/handle/n4zt38.
Повний текст джерелаMota, Ana Catarina Candeias. "Real-time droplet monitoring for digital Polymerase Chain Reaction in microfluidic chip." Master's thesis, 2020. http://hdl.handle.net/10362/117487.
Повний текст джерелаAs técnicas actuais usadas no diagnóstico de cancro, geralmente dependem da recolha de tecido tumoral, envolvendo processos invasivos para o paciente. O DNA tumoral circu lante (ctDNA) surge como alternativa para a detecção e monitorização do cancro, podendo ser extraído através de amostras de sangue. A reação em cadeia da polimerase de modo digital (dPCR) é uma técnica rápida e sensível para amplificação de DNA, adequado para baixas concentrações de DNA, como o ctDNA. Os avanços na microfluídica permitem a partição das amostras de PCR em gotas com base em emulsões de água em óleo, de modo que a amplificação por PCR ocorra dentro de cada gota. Deste modo, a reação de PCR é um processo bem controlado com baixa probabilidade de contaminação, permitindo uma análise de alto rendimento. Este trabalho teve como objetivo o desenho e a fabricação de um dispositivo de micro fluídica capaz de produzir um elevado número de gotas uniformes, cujos volumes se encontram na gama dos nanolitros, com frequência constante. Para tal, foi desenvolvido um dispositivo para geração de gotas em polidimetilsiloxano (PDMS), através de técnicas de fotolitografia e litografia suave, tendo sido testado com diversas taxas de fluxos entre óleo / água. Posteriormente, as gotas geradas foram caracterizadas em relação ao seu ta manho, velocidade e frequência através do software de análise de vídeo Bonsai. Diversos testes em diferentes dispositivos foram realizados de modo a avaliar a reprodutibilidade do dispositivo. Por último, o gerador de gotas foi incorporado com desenho da serpentina, permitindo que os ciclos de PCR ocorram em fluxo contínuo. Os estudos realizados revelaram que foi possível gerar gotas com raios entre 22-99 µm, e coeficiente de variação inferior a 10%. Os volumes correspondentes variaram entre 90 pL 4.18 nL. Além disso, as velocidades obtidas situaram-se entre 0.05 mm/s-7.62 mm/s com frequência de geração de gotas de 2-50 Hz. Relativamente à monitorização das gotas, os resultados dos workflows desenvolvidos revelaram similaridade com os resultados obtidos através de um software amplamente utilizado para estes fins, com a vantagem de permitir a análise em tempo real para uma amostra maior de resultados.
Chen, An-Te, and 陳安得. "DNA Extraction and Real-time Polymerase Chain Reaction on Digital Microfluidic Chip." Thesis, 2016. http://ndltd.ncl.edu.tw/handle/77633924624998769116.
Повний текст джерела國立臺灣大學
機械工程學研究所
105
This thesis reports the implementation of DNA extraction and real-time PCR (qPCR) on a digital microfluidic (DMF) device. We aim to develop a personalized point-of-care device for molecular diagnosis from human whole blood and commercial reagents kits driven by electrowetting-on-dielectric (EWOD) on a DMF device for DNA extraction and qPCR. The results from on-chip DNA extraction protocols were validated and quantified. In comparison wiht the traditional DNA extraction procedures in tubes, our on-chip extraction starting from a 100 nL whole blood obtained 52.8 % DNA concenttion at 4.45 ng/µL, required 25 % reaction time (from 120 min to 30 min), consumed 0.05 % volume of the entire reagents (from 4222
Chang, Yi-Hsien, and 張儀賢. "Digital Microfluidic Chips with Low Operation Voltages for Polymerase Chain Reaction pplications." Thesis, 2005. http://ndltd.ncl.edu.tw/handle/53803224655745668820.
Повний текст джерела國立成功大學
工程科學系碩博士班
93
This study designs and fabricates a digital microfluidic platform with a low operation voltage by utilizing electro-wetting-on-dielectric (EWOD) effect. All the microfluidic movements are realized bet ween two parallel plates on this platform. The digital microfluidics chips (DMC) could make liquids digitalized, allowing for the transportation of droplets without moving micromechanical components in the device. Most fluidic operations such as sample transporting, creation, cutting and merging can be carried out on the chips using discrete droplets. The DMC does not require pumps, mixers and valves as in traditional microfluidic systems. The droplet motion can be manipulated by using surface tension gradient generated by EWOD, which could control of the wettability of liquids on dielectric thin film surfaces while electric potential is applied. This study tested several high K materials such as BST (Ba0.5Sr0.5TiO3), Ta2O5, Si3N4, Parylene and silicon-dioxide (SiO2). By using the Young-Lippmann equation, reducing the operating voltage of EWOD could be realized. Meanwhile, the hysteresis effect of contact angles between the droplet and the Teflon surface could be reduced by filling silicone oil into the parallel open channel. Therefore, the four fundamental microfluidics operations, transport, creation, cutting, and merging, can be performed with a lower voltage of 12 V. Polymerase chain reaction (PCR) is a popular technology in molecular biology. A micro temperature sensor and micro heaters have been integrated into the EWOD chip to allow for PCR operation, which is dependent on three precise temperature steps. Utilizing the characteristics of a temperature with a constant TCR (Temperature Coefficient of Resistance), this study designed a closed-loop temperature controller. The micro temperature sensor, micro heaters, and smaller reagent volume allow for a heating rate of 38.0 degrees per second and a cooling rate of 7.9 degrees per second, thus enabling for faster completion of the PCR cycles. The micro temperature sensor, micro heaters are controlled by a commercial programmable chip (AT90S8535). This chip successfully achieved 25 thermo cycles PCR for the genes of Dengue II Virus cDNA in 55 minutes. Utilizing micro-electro-mechanical-system (MEMS) fabrication technology, the digital microfluidics operation system and micro temperature controller could be successfully integrated into a cheap and biocompatible glass substrate to allow automated rapid infectious disease detection. By using EWOD effect, the chip could make liquids digitalized, permitting droplets to be quickly and precisely transported and mixed without any moving components. This study also designed a new hydrophilic-hydrophobic interface to induce the surface tension gradient to transport the mixed droplets into the reaction chamber. The Dengue II Virus cDNA polymerase chain reaction was also performed on the developed chip which was used to detect microorganism genes by amplifying the target sequence (511bps) with PCR. This study confirmed the feasibility of the PCR with the digital microfluidics operation. The integrated DMC has several advantages for infectious disease detection including (1) low voltage operation DMC, (2) 70% less sample and reagent consumption (3) 50% shorter detection time due to faster thermal cycling, and (4) high accuracy and reproducibility due to a simple and reliable fabrication process. The study successfully applied the concept of Digital Micro Fluidics to the Lab-on-a-chip. The developed microchip will be of great benefit in the future genetic analysis and infectious disease detection.
Chiang, Tsai-Jung, and 蔣采蓉. "Polymerase Chain Reaction of DNA by Joule Heating on an EWOD-Based Digital Microfluidic Platform." Thesis, 2011. http://ndltd.ncl.edu.tw/handle/52766787872652536373.
Повний текст джерела國立交通大學
材料科學與工程學系奈米科技碩博士班
99
We investigated joule heating to treat a small amount of biochemical sample driven by electrowetting-on-dielectric (EWOD) on a digital microfluidic platform. For the decreased reagent volume and reduced device size, the heating and cooling rates are thus increased. With the heating and driving abilities, we utilize this platform to realize polymerase chain reaction of DNA with fast heating and cooling rates. On the contrary, conventional equipments usually give lower heating and cooling rates because large amounts of biochemical samples and reagents are required. The reported digital microfluidic system with the joule heating ability would provide an approach to solve the heating/cooling rate issues by reducing the reaction volume from conventional 50 ?愮 to 3 ?愮 at a heating rate of 2.46 oC/s and a cooling rate of 6.94 oC/s The system consumes a much lower power 8.5 x 10-3 W than that consumed in traditional PCR machines. In the experiment, the external voltage at low frequency (1 kHz) can drive droplet by EWOD without temperature change. At high frequency (higher than 100 kHz), the AC electric field heats the droplet and causes an 80 oC temperature change to the temperature of 105.6 oC. The temperature of the tested droplet rises when the external voltage increases. In addition, the heating is also frequency-related. Therefore, this system can control the temperature and position of the droplet for realizing the polymerase chain reaction- by tuning the amplitude and frequency of the applied voltage.
"An investigation into the determination of relative chromosome dosage by digital PCR." 2009. http://library.cuhk.edu.hk/record=b5896560.
Повний текст джерелаThesis (M.Phil.)--Chinese University of Hong Kong, 2009.
Includes bibliographical references (leaves 133-150).
Abstract also in Chinese.
ABSTRACT --- p.i
摘要 --- p.iii
ACKNOWLEDGEMENTS --- p.iv
CONTRIBUTORS --- p.vi
TABLE OF CONTENTS --- p.vii
LIST OF TABLES --- p.x
LIST OF FIGURES --- p.xi
LIST OF ABBREVIATIONS --- p.xiii
Chapter SECTION I: --- BACKGROUND --- p.1
Chapter CHAPTER 1: --- PRENATAL DIAGNOSIS OF FETAL TRISOMY 21 --- p.2
Chapter 1.1 --- Down syndrome --- p.2
Chapter 1.2 --- Current methods of prenatal diagnosis of fetal trisomy 21 --- p.3
Chapter 1.2.1 --- Non-invasive procedures --- p.3
Chapter 1.2.2 --- Invasive procedures --- p.5
Chapter 1.3 --- Alternative methods for the prenatal diagnosis of fetal trisomy 21 --- p.7
Chapter CHAPTER 2: --- CELL-FREE FETAL NUCLEIC ACIDS IN MATERNAL PLASMA --- p.13
Chapter 2.1 --- Circulating fetal cells --- p.15
Chapter 2.2 --- Circulating cell-free fetal nucleic acids --- p.15
Chapter 2.3 --- Diagnostic applications of cell-free fetal nucleic acids in maternal plasma --- p.17
Chapter 2.4 --- Digital relative chromosome dosage approach --- p.20
Chapter 2.5 --- Validation of digital RCD approach on artificial DNA mixtures --- p.22
Chapter SECTION II --- : MATERIALS AND METHODS --- p.25
Chapter CHAPTER 3: --- QUANTITATIVE ANALYSIS OF NUCLEIC ACIDS --- p.26
Chapter 3.1 --- Subject recruitment and sample collection --- p.26
Chapter 3.2 --- Sample processing --- p.26
Chapter 3.3 --- Nucleic acid extraction --- p.27
Chapter 3.3.1 --- Extraction of DNA from placental tissues --- p.27
Chapter 3.3.2 --- Extraction of DNA from maternal blood cells --- p.27
Chapter 3.4 --- Matrix-assisted laser desorption/ionisation time-of-flight mass spectrometry (MALDI-TOF MS) --- p.28
Chapter 3.5 --- Paralogous sequence assays optimisation workflow --- p.31
Chapter 3.5.1 --- Monoplex paralogous sequence assays --- p.31
Chapter 3.5.2 --- Multiplex paralogous sequence assay --- p.38
Chapter 3.6 --- Digital PCR --- p.42
Chapter 3.6.1 --- Principle --- p.42
Chapter 3.6.2 --- Digital multiplex paralogous sequence assay --- p.42
Chapter 3.7 --- Statistical analysis --- p.46
Chapter 3.7.1 --- Disease classification of samples --- p.46
Chapter 3.7.2 --- Poisson distribution --- p.46
Chapter 3.7.3 --- Data analysis --- p.48
Chapter 3.7.4 --- Sequential probability ratio test (SPRT) analysis --- p.49
Chapter SECTION III: --- ASSAY DEVELOPMENT --- p.53
Chapter CHAPTER 4: --- TESTING OF ASSAY SPECIFICITY WITH CORIELL CELL LINES --- p.54
Chapter 4.1 --- Coriell cell lines --- p.54
Chapter 4.2 --- Specificity of initial PCR primers --- p.56
Chapter 4.2.1 --- Principle --- p.56
Chapter 4.2.2 --- Materials and methods --- p.56
Chapter 4.2.3 --- Results --- p.60
Chapter 4.2.4 --- Conclusion --- p.63
Chapter 4.3 --- Specificity of the iPLEX® Gold extension primers --- p.63
Chapter 4.3.1 --- Principle --- p.63
Chapter 4.3.2 --- Materials and methods --- p.64
Chapter 4.3.3 --- Results --- p.65
Chapter 4.4 --- Further analysis on the specificity of PV2107a initial PCR primers --- p.67
Chapter 4.5 --- Conclusion --- p.71
Chapter CHAPTER 5: --- ASSAY OPTIMISATION --- p.72
Chapter 5.1 --- Introduction --- p.72
Chapter 5.2 --- Optimisation of initial PCRs with AmpliTaq Gold® DNA polymerase followed by homogeneous MassEXTEN´DёØ (hME) assays (Sequenom) --- p.72
Chapter 5.2.1 --- Optimisation of initial PCR reactions --- p.72
Chapter 5.2.2 --- Principle of homogeneous MassEXTEN´DёØ assays (Sequenom)… --- p.75
Chapter 5.2.3 --- Homogeneous MassEXTEN´DёØ assays (Sequenom) on euploid and T21 samples --- p.76
Chapter 5.3 --- Assay selection by iPLEX® Gold single base primer extension reactions (Sequenom) --- p.82
Chapter 5.4 --- Optimisation of multiplex PCR with AmpliTaq Gold® DNA polymerase --- p.88
Chapter 5.5 --- Optimisation of multiplex iPLEX® Gold single base primer extension reaction --- p.93
Chapter 5.6 --- Single molecule detection test for the multiplex paralogous sequence assays … --- p.103
Chapter SECTION IV: --- ANALYSIS OF CLINICAL SAMPLES --- p.107
Chapter CHAPTER 6: --- DISEASE CLASSIFICATION OF EUPLOID AND TRISOMY SAMPLES WITH MULTIPLEX PARALOGOUS SEQUENCE ASSAY --- p.108
Chapter 6.1 --- Introduction --- p.108
Chapter 6.2 --- Materials and methods --- p.109
Chapter 6.2.1 --- Sample collection --- p.109
Chapter 6.2.2 --- Experimental design --- p.110
Chapter 6.3 --- Results --- p.111
Chapter 6.4 --- Discussion --- p.114
Chapter SECTION V: --- CONCLUDING REMARKS --- p.122
Chapter CHAPTER 7: --- CONCLUSION AND FUTURE PERSPECTIVES --- p.123
Chapter 7.1 --- Conclusion --- p.123
Chapter 7.2 --- Future perspectives --- p.124
Appendix 1 --- p.126
Appendix II --- p.127
REFERENCE --- p.133
Su, Ying-Tu, and 蘇英圖. "Application of MicroEmusification:Emulsion Polymerase Chain Reaction." Thesis, 2009. http://ndltd.ncl.edu.tw/handle/51940901593932821213.
Повний текст джерела國立臺灣海洋大學
機械與機電工程學系
97
In this thesis, soft lithography technique was used to produce a micro emulsion chip. This chip can handle tiny volume of DNA solution for polymerase chain reation (ePCR). The DNA solution is dispersed in the emulsified droplets in a continuous oil phase, and the droplets become reaction space for PCR. As a result, the consistency of the multiple types of DNA templates existing in the original reaction space can be improved by emulsification process. Also, it will reduce competitive sequence interference and improve DNA magnification. Differing from usual micro emulsification chips, this study focuses on the technical problems involved in dealing with tiny and expensive DNA solution in order to avoid non-uniform droplets in the transient of droplet generation. The main ideas are as follows: (1) Let water first go through transient process and produce water droplets; (2) Use water and bubble to place DNA solution in between, and pass it through emulsification hydrodynamic focusing channel; (3) Use pneumatic membrane valves control the stop and flow of water and DNAsolution. Characterization of the emulsification chips includes droplet sizes, uniformity of droplets and hot-resistance of water-in-oil droplets. The smallest droplet diameter is less than 10 μm and the coefficient of variation is about 2%. The resulting electrophoretograms of both single or multiple templates’ PCR demonstrate that ePCR can improve the precision of amplified DNA segments. It is suggested from the experiment results that ePCR was proven to be efficient and could be used in further genetic research Key word: ePCR, DNA template, transient process, tiny-volume micro-emulsification
Gi, Kao Li, and 高麗姬. "ABO Genotyping for Mutiplex Polymerase Chain Reaction." Thesis, 1999. http://ndltd.ncl.edu.tw/handle/82466547226309854097.
Повний текст джерела中央警察大學
刑事警察研究所
87
Abstract ABO blood group is clinically the most important blood group system in transfusion medicine including fours common different phenotypes . It has been a valuable tool in transfusion medicine , physical anthropology , disputed parentage testing , human identification , and in forensic analysis . Historically , forensic and clinical labatories utilize serological techniques to identify ABO blood type . The testing of forensic samples for ABO types using serological techniques has several drawbacks . We has simplified the identification of ABO types by taking advantage of previouly reported ABO DNA sequence difference . The use of single strand conformation polymorphism(SSCP)can separate sequence polymorphism than differ by only one base . We has investigated in 260 Chinese donors by multiplexed Polymerase Chain Reaction . In this study the ABO alleles from exon 6 , producing a 212/213 bp fragment , and exon 7, which produces a fragment of 272 bp , were separated by non-denaturing polyacrylamide gels . The two exons were amplified in a single reaction that produces similar quantities of DNA for both exons . However , in 260 SSCP patterens we can clustered 9 into different distinct and discernible groups . A concordance rate of 98.5 ﹪(256/260 samples)observed between the actural genotype and the serologically-based predicted genotype . These results indicate that the assay provides a rapid , accurate , and simple method for ABO genotyping that serves as a useful supplement to standard serological ABO typing .
Chen, Jhao-Rong, and 陳昭榮. "Micro Rayleigh-Bénard polymerase chain reaction system." Thesis, 2005. http://ndltd.ncl.edu.tw/handle/05888212591754965876.
Повний текст джерела國立清華大學
工程與系統科學系
93
Polymerase chain reaction (PCR) is a molecular biological method for the in vitro amplification of nucleic acid molecule. In this thesis, the research object was to design a micro PCR system which involved a Rayleigh-Bénard convection PCR chip, measurement circuits, and temperature control circuits. Rayleigh-Bénard convection PCR chip was easy to be fabricated, and the sample solution in it can transit its temperature immediately. Thus, the faster the speed of flow is, the higher heating and cooling rate is. Rayleigh-Bénard convection PCR chip can be divided into two parts. First part is a chip with micro heaters and micro temperature sensors. Second part is PDMS reaction chamber designed by Rayleigh-Bénard convection theory. Temperature control system is designed to keep temperature on the top and bottom chips by Pulse width modulation control. Analog temperature signal is precisely measured by circuits. Then the signal is processed by an 8051 single chip, and the output power of micro heaters is controlled to adjust the temperature of top and down plates. Finally, agarose gel electrophoresis is used to verify the practicability of Rayleigh-Bénard convection PCR system. By comparing with the PCR experiments done by the commercial PCR machine and our chips, our chips can plenty decrease the reaction time. And By comparing with the simulation and PCR experiments of different designed sizes, users can use setted parameters and CFD results to do optimal designs and then decrease the total reaction time in the future.
Tao, Yo-Shen, and 刁宥升. "Design Of Polymerase Chain Reaction System Controller." Thesis, 2002. http://ndltd.ncl.edu.tw/handle/76827215729605694932.
Повний текст джерела國立暨南國際大學
電機工程學系
90
DNA analysis is a crucial process for biotechnology. Polymerase chain reaction (PCR) is an imperative step for performing DNA analysis. PCR can proliferate a small quantity of DNA sample to a large quantity enough for DNA analysis in a short time. Our goal is to develop a PCR system chip by integrating standard CMOS process and MEMS process. In this thesis, an 8-bit 4-level pipelined programmable micro-controller for temperature control during PCR process is designed by HDL description and implemented by using field programmable gate array (FPGA). The micro-controller includes modules of data path and control unit. Verilog hardware description language is used to model the modules. The individual module is functionally verified by Max Plus II of Altera and integrated into an 8-bit micro-controller. The micro-controller is realized by a FPGA chip, and then micro program for the temperature control of PCR is downloaded into the ROM in FPGA for system test. The micro-controller proposed in this thesis is very simple and easy to implement. It is suitable for general low speed applications. In order to easily integrate the micro-controller into other applications, we will try to parameterize its design variables for adjusting its hardware structure in future works.
Gao, Li-Ji, and 高麗姬. "ABO Genotyping for Multiplex Polymerase Chain Reaction." Thesis, 1998. http://ndltd.ncl.edu.tw/handle/7433nn.
Повний текст джерела中央警察大學
刑事警察研究所
87
ABO blood group is clinically the most important blood group system in transfusion medicine including fours common different phenotypes . It has been a valuable tool in transfusion medicine , physical anthropology , disputed parentage testing , human identification , and in forensic analysis . Historically , forensic and clinical labatories utilize serological techniques to identify ABO blood type . The testing of forensic samples for ABO types using serological techniques has several drawbacks . We has simplified the identification of ABO types by taking advantage ofpreviouly reported ABO DNA sequence difference . The use of single strand conformation polymorphism ( SSCP ) can separate sequence polymorphism than differ by only one base . We has investigated in 260 Chinese donors by multiplexed Polymerase Chain Reaction . In this study the ABO alleles from exon 6 , producing a 212/213 bp fragment , and exon 7, which produces a fragment of 272 bp, were separated by non-denaturing polyacrylamide gels. The two exons were amplified in a single reaction that produces similar quantities of DNA for both exons . However , in 260 SSCP patterens we can clustered 9 into different distinct and discernible groups . A concordance rate of 98.5 % (256/260 samples) observed between the actural genotype and the serologically-based predicted genotype . These results indicate that the assay provides a rapid , accurate , and simple method for ABO genotyping that serves as a useful supplement to standard serological ABO typing.