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Příprava vzorků pro analýzu DNA v potravinách rostlinného původu / Sample preparation for DNA analysis from foods of plant originSilná, Renata January 2018 (has links)
The isolation of high quality DNA is nessecary for many molecular biology applications. However, plant DNA contains high amonts of polysaccharides, polyphenols and various secondary metabolites, which decrease yield and quality of isolated DNA. The aim of this study was preparation of samples and different food matrices for DNA isolation DNA by magnetic particles. It was about 5 species of vegetable and 10 species of processed plant food. Homogenization of samples was performed in CTAB buffer. Isolation of plant DNA was performed by magnetic particles covered with carboxyl groups. All DNAs were isolated in conventional PCR qualities using primers for 700 bp amplicons, in the case of heat processed products for 220 bp ampilicons and for real time PCR. The efficiancy of separation of magnetic particles with DNA by magnetic separator and magnetic needle was compared. It was find out that DNA of higher purity was isolated using magnetic needle. The micromethod of isolation of plant DNA from homogenates with CTAB with magnetic particles is suitable for different processed food.
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Využití různých metod izolace DNA baktérií mléčného kvašení v molekulárně biologických metodách / Using different methods of DNA isolation of lactic acid bacteria in molecular biological methodsChvalkovská, Eva January 2019 (has links)
This thesis focused on the probiotic bacteria, DNA isolated from these bacteria by three different methods and the effect of isolation on DNA identification using molecular biological methods. Probiotic bacteria are an important part of human intestinal tract. They have an important role in the function of the immune system due to adhesion to the mucosa of the intestinal flora. They create a inhostile environment for pathogens. Probiotic bacteria are commonly taken in the food like dairy products or food supplements. However, overuse of antibiotics is at risk of passing on the intrinsic resistance that probiotic bacteria have to the pathogenic bacteria. The intrinsic resistence they have to maintain the natural homeostasis of the intestinal tract. It is important to effectively identify risky probiotic bacteria that have the ability to transmit resistance to eliminate their presence in food and dietary supplements. Three methods of DNA isolation like phenol extraction method, magnetic particle isolation and commercial kit isolation were used in the experimental part. DNA was isolated from three dietary supplements, namely Biopron 9 premium, Linex forte and GS Lactobacily forte 21. The purity and concentration of the isolated DNA was detected spectrophotometrically. The presence of individual DNA strains in dietary supplements was confirmed by real-time polymerase chain reaction. The best method of isolation in terms of purity and concentration of isolated DNA was evaluated by RT-PCR and spectrophotometry using a commercial kit isolation method.
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Izolace DNA z probiotických druhů baktérií mléčného kvašení v potravinových doplňcích / DNA isolation from probiotic lactic acid bacteria in food additivesTvrdíková, Jana January 2009 (has links)
In this work the functionalised magnetic particles were tested with streptavidin to selective DNA isolation. The method of selective DNA isolation was tested by using DNA probiotic strain Lactobacillus paracasei subsp. paracasei CCDM 211/06. A test was done on the biotinyl oligonucleotic particles, which was immobilised by containing streptavidin and it was used like a DNA probe for isolation complementary DNA chain by means of DNA/DNA hybridization. The primer R 5´ bio and the biotinyl denatured specific PCR product were tested for species Lb. paracasei as a DNA probe. These following experimental conditions were optimized for selective DNA isolation: temperature and time of hybridization, amount of DNA and the release of DNA from microspheres. Isolation of DNA was verified by PCR with specific generic primers. The specific generic PCR product was amplified in extent 250 bp, which was detected by using electrophoresis in agarose gel. This optimized method was successfully used in selective isolation of DNA Lactobacillus from a complementary sample of supplementary food (BIFI pangamin).
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Izolace DNA z probiotických výrobků s využitím pevných nosičů / Isolation of DNA from probitic products using solid carriersBonczek, Ondřej January 2011 (has links)
Microbial DNA was isolated from lysed cells of Lactobacillus genus in probiotic products. Reversible adsorption DNA on the surface of carboxyl coated nonporous poly(2-hydroxyethyl methacrylate-co-glycidyl methacrylate) (P(HEMA-co-GMA)) magnetic particles and silicagel coated manganase Perovskite nanoparticles. DNA was adsorbed on the surface of the particles in the presence of 16 % poly(ethylenglycol) (PEG 6000) and 2 M sodium chloride (NaCl) concentrations. The adsorbed DNA was released from particles by low ionic strength TE buffer (pH= 8.0). The quality of isolated DNA was checked by spectrofotometric measurement and PCR amplification. DNA samples isolated using magnetic particles and phenol extraction method (control method) were PCR-ready. The DNA isolated from lysed cells of probiotic products was quantificated in real-time qPCR.
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Selektivní izolace bakterií rodu Bifidobacterium z potravin / Selective isolation of the genus Bifidobacterium bacteria from foodsMizerovská, Lucie January 2012 (has links)
Probiotic lactic acid bacteria (LAB) are very often used in food procesing industry, such as milk products, cheese and fermentsd salami production in nova days. In diploma thesis were tested symbiotic food supplements from different producers. Bacterial DNA was isolated from crude cell lysates of six food suplements by magnetic particles P(HEMA-co-GMA). PCR-ready DNAs were isolated. from all products The detection of Bifidobacterium bacteria identified by PCR was in agreement with those declared by the manufacturers. Magnetic particles with immobilized antibodies against Bifidobacterium were used in the next part of thesis. These particles were used for the isolation of target cells from two products with cell identification by genus specific PCR.
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Studium reverzibilní adsorpce nukleových kyselin na pevných nosičích / Study of Reversible Adsorption of Nucleic Acids on Solid SurfacesTrachtová, Štěpánka January 2011 (has links)
Magnetically driven separation techniques using magnetic solid carriers are one of modern methods to speed up and facilitate the previously used separation and purification procedures. The use of magnetic particles in biology imposes strict requirements on physical, and chemical properties of the particles, including low toxicity, biocompatibility and non-interference with the chemical environment in diagnostics. The aim of this study was to evaluate carboxyl-functionalised magnetic non-porous P(HEMA-co-GMA), P(HEMA-co-EDMA), PGMA, silica-coated lanthanum manganese peroskvite La0.75Sr0.25MnO3 and thermosensitive poly(N-isopropylacrylamide) microspheres – P(NIPAAm) for DNA isolation from different types of complex food and environmental samples containing PCR inhibitors. The solid-phase reversible immobilisation (SPRI) of nucleid acids on microsphere surface and the release of adsorbed DNA were optimised. DNA from real samples (milk products and probiotic food suplements, mouse faeces) was apparently adsorbed on solid particles from the aqueous phase system composed of 16% PEG 6000 and 2M NaCl. The conditions of the subsequent release absorbed DNA to the elution buffer (pH of elution buffer, temperature and time of elution) were optimized. The quality of eluted DNA and the presence of target DNA were examined by PCR and q-PCR using domain-specific Bacteria and genus-specific Lactobacillus primer set. Real-time PCR was used for an estimation of the PCR interference by comparing the amplification efficiencies of purified DNA containing solid nanoparticles with the DNA standards free of any nanoparticles
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Využití magnetických částic při izolaci DNA z vybraných probiotických výrobků pro děti / The application of magnetic particle for DNA isolation from selekted probiotic products for childrenVozárová, Petra January 2017 (has links)
In the food industry, it is important to correctly identify the species of bacteria and thier properties so that they can be used as a probiotic in dietary supplements. This is performed using DNA diagnostics. In the experimental part, the DNA from four probiotic dietary supplements for children was isolated. Magnetic particles P(HEMA-co-GMA) were tested for isolation. Isolated DNA was amplified by PCR and the presence of DNA of genus Lactobacillus, Bifidobacterium and Bacillus was demonstrated in the products according to the data declared by the manufacturer. The presence of species L.acidophilus, B.animalis in accordance with the data on the product has been demonstrated by PCR with species specific primers. Using PCR, the presence of L.casei, which was declared by the manufacturer, has not been proven in one product at given experimental conditions.
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Tau protein, biomarker Alzheimerovy choroby: in vitro fosforylace a charakterizace tau reaktivních protilátek / Tau protein, a biomarker of Alzheimer's disease: in vitro phosphorylation and tau-reactive antibodies characterizationHromádková, Lenka January 2018 (has links)
Tau protein, a microtubule-associated protein localized in axonal projections of neurons, is a key molecule in the pathology of Alzheimer's disease (AD), the most common cause of dementia in the elderly population. Tau belongs to the group of natively unfolded proteins without globular structure and is prone to numerous posttranslational modifications (PTMs). Under pathological conditions, abnormal PTMs and misfolding of tau protein occurs and leads to oligomerization and aggregation into paired helical filaments forming neurofibrillary tangles, the histopathological hallmark of AD. Currently available drugs applied in AD treatment can only slow the disease progression and those, which halt the AD-specific neurodegenerative processes, are still missing. Very promising and evolving therapeutic approach is immunotherapy, and even immunomodulation by administration of intravenous immunoglobulin (IVIG) products, a reservoir of natural antibodies from the plasma of healthy donors, has been already tested. The discovery of naturally occurring antibodies directed to tau (nTau-Abs) in body fluids of both AD and healthy subjects and their presence in IVIG begin the investigation of their therapeutic potential. Considering a wide range of possible modifications of tau and of various tau species (oligomers,...
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[pt] MICROSCOPIA MAGNÉTICA DE VARREDURA UTILIZANDO SENSORES DE EFEITO HALL COM POLARIZAÇÃO DE CORRENTE PULSADA / [en] SCANNING MAGNETIC MICROSCOPY USING HALL EFFECT SENSORS BIASED WITH PULSED CURRENTLANNA ISABELY MORAIS SINIMBU 19 October 2023 (has links)
[pt] A pesquisa de caracterização de materiais é fundamental para o
desenvolvimento de diversas tecnologias. No campo de estudo de amostras
anisotrópicas como o caso encontrado no paleomagnetismo, é importante estudar
a caracterização magnética de minerais rochosos para compreender o campo
geomagnético. A caracterização de amostras anisotrópicas, como rochas, tem
aplicações na datação do tempo geológico e na caracterização de minérios que
sofrem mudanças magnéticas em temperaturas superiores à de Curie. Assim, é
necessário conhecer as propriedades físicas desses materiais em relação ao
comportamento magnético. Existem várias instrumentações usados neste estudo,
como magnetômetro utilizando sensores Superconducting Quantum Interference
Device (SQUID), magnetômetro de Amostra Vibrante (VSM), Magnetic Property
Measurement System (MPMS) e os Microscópios Magnéticos de Varreduras
(MMV) entre outros. Entretanto, o MMV permite mapear o comportamento
magnético do material realizando desta forma uma caracterização magnética local
da amostra. Nesse contexto, foi proposta uma alternativa de leitura usando a
técnica Delta Mode da empresa Keithley no MMV. O objetivo é eliminar os efeitos
termoelétricos, realizar medições com baixo ruído de tensão e obter uma
alternativa para a leitura do campo magnético induzido no MMV. No
desenvolvimento do trabalho, foram utilizados sensores de efeito Hall do modelo
HQ-811 em uma configuração gradiométrica, juntamente com uma fonte de
corrente (Keithley, modelo 6220) e um Nanovoltímetro (Keithley, modelo
2182A). Os resultados iniciais da calibração revelaram um desvio de cerca de 6 por cento
nas medições. Com base nessa calibração, nosso objetivo final é empregar a
técnica Delta Mode no MMV para obter as curvas de magnetização das amostras,
mesmo em situações desafiadoras de sobreposição de sinais de campo magnético
induzido. A metodologia adotada envolve o uso de partículas magnéticas de óxido
de ferro em escala nano e micro, posicionadas em três cavidades cilíndricas (A1,
A2 e A3) de aproximadamente 800 micrômetros de diâmetro com 800 micrômetros de profundidade,
dentro do mesmo porta-amostra. Durante o experimento, um dos desafios foi lidar
com a sobreposição de sinais quando aplicado um campo magnético perpendicular
ao plano do porta-amostra. Para determinar o valor de magnetização de cada
amostra, utilizamos um modelo teórico de um cilindro de corrente devido ao
formato das cavidades onde as amostras foram depositadas. Os valores de
magnetizações encontrados para diferentes espaçamentos de posicionamento das
amostras foram em média de AM1= 62,59 Am(2)
/kg, AM2= 13,14 Am(2)
/kg e AM3=
10,13 Am(2)
/kg para um campo de 0,5 T, onde apresentaram reprodutividade para
diferentes situações. / [en] Materials characterization research is fundamental for the development of various technologies. In the field of anisotropic sample studies, such as in paleomagnetism, it is important to investigate the magnetic characterization of rock minerals to understand the geomagnetic field. Characterizing anisotropic samples, such as rocks, has applications in geological time dating and the characterization of ores that undergo magnetic changes at temperatures higher than the Curie temperature. Therefore, it is necessary to understand the physical properties of these materials in relation to their magnetic behavior. Various instrumentation techniques are used in this study, including magnetometers using Superconducting Quantum Interference Device (SQUID) sensors, Vibrating Sample Magnetometer (VSM), Magnetic Property Measurement System (MPMS), and Magnetic Scanning Microscopes (MMV), among others. However, MMV allows for mapping the magnetic behavior of the material, thus providing a local magnetic characterization of the sample. In this context, an alternative reading technique using the Delta Mode of the Keithley company was proposed for MMV. The objective is to eliminate thermoelectric effects, perform low-noise voltage measurements, and provide an alternative for reading the induced magnetic field in MMV. In the course of this work, Hall effect sensors of the HQ-811 model in a gradiometric configuration, along with a current source (Keithley, model 6220), and a Nanovoltmeter (Keithley, model 2182A) were used. The initial calibration results revealed a deviation of about 6 percent in the measurements. Based on this calibration, our ultimate goal is to employ the Delta Mode technique in the MMV to obtain the magnetization curves of the samples, even in challenging situations of overlapping induced magnetic field signals. The adopted methodology involves the use of nano- and micro-scale iron oxide magnetic particles positioned in three cylindrical cavities (A1, A2, and A3) of approximately 800 micrometers in diameter with 800 micrometers in depth, within the same sample holder. During the experiment, one of the challenges was dealing with signal overlap when a magnetic field was applied perpendicular to the plane of the sample holder. To determine the magnetization value of each sample, a theoretical model of a current cylinder was used due to the shape of the cavities where the samples were deposited. The magnetization values found for different sample positioning spacings were on average AM1 = 62.59 Am(2)/kg, A(2) = 13.14 Am(2)/kg, and A3 = 10.13 Am(2)/kg for a magnetic field of 0.5 T, which showed reproducibility for different situations.
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Initial Attachment of Pseudomonas Aeruginosa on Modified Polycardonal CoatingsSharma, Lohit, sharma January 2016 (has links)
No description available.
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