Optimization of sample preparation for next-generation sequencing with Illumina Genome Analyzer II

The growing interest for sequencing with higher throughput in the last decade has led to the development of new sequencing applications. This thesis concentrates on optimizing DNA library preparation for Illumina Genome Analyzer II sequencer. The library preparation steps that were optimized include fragmentation, PCR purification and quantification. DNA fragmentation was performed with focused sonication in different concentrations and durations. Two column based PCR purification method, gel matrix method and magnetic bead based method were compared. Quantitative PCR and gel electrophoresis in a chip were compared for DNA quantification. The magnetic bead purification was found to be the most efficient and flexible purification method. The fragmentation protocol was changed to produce longer fragments to be compatible with longer sequencing reads. Quantitative PCR correlates better with the cluster number and should thus be considered to be the default quantification method for sequencing. As a result of this study more data have been acquired from sequencing with lower costs and troubleshooting has become easier as qualification steps have been added to the protocol. New sequencing instruments and applications will create a demand for further optimizations in future.

Escape of high mass ions due to initial thermal energy and its implications for axially symmetric RF ion trap mass analyzer design

This paper investigates the loss of high mass ions due to their initial thermal energy in ion trap mass analyzers. It provides an analytical expression for estimating the percentage loss of ions of a given mass at a particular temperature, in a trap operating under a predetermined set of conditions. The expression we developed can be used to study the loss of ions due to its initial thermal energy in traps which have nonlinear fields as well as those which have linear fields. The expression for the percentage of ions lost is shown to be a function of the temperature of the ensemble of ions, ion mass and ion escape velocity. An analytical expression for the escape velocity has also been derived in terms of the trapping field, drive frequency and ion mass. Because the trapping field is determined by trap design parameters and operating conditions, it has been possible to study the influence of these parameters on ion loss. The parameters investigated include ion temperature, magnitude of the initial potential applied to the ring electrode (which determines the low mass cut-off), trap size, dimensions of apertures in the endcap electrodes and RF drive frequency. Our studies demonstrate that ion loss due to initial thermal energy increases with increase in mass and that, in the traps investigated, ion escape occurs in the radial direction. Reduction in the loss of high mass ions is favoured by lower ion temperatures, increasing low mass cut-off, increasing trap size, and higher RF drive frequencies. However, dimensions of the apertures in the endcap electrodes do not influence ion loss in the range of aperture sizes considered. (C) 2010 Elsevier B.V. All rights reserved.

Vibration spectrum analyzer using stretched membranes of polymer piezoelectrics for sensor networks

The key problem tackled in this paper is the development of a stand-alone self-powered sensor to directly sense the spectrum of mechanical vibrations. Such a sensor could be deployed in wide area sensor networks to monitor structural vibrations of large machines (e. g. aircrafts) and initiate corrective action if the structure approaches resonance. In this paper, we study the feasibility of using stretched membranes of polymer piezoelectric polyvinlidene fluoride for low-frequency vibration spectrum sensing. We design and evaluate a low-frequency vibration spectrum sensor that accepts an incoming vibration and directly provides the spectrum of the vibration as the output.

A radial differential mobility analyzer for the size-classification of gas-phase synthesized nanoparticles at low pressures

Differential mobility analyzers (DMAs) are commonly used to generate monodisperse nanoparticle aerosols. Commercial DMAs operate at quasi-atmospheric pressures and are therefore not designed to be vacuum-tight. In certain particle synthesis methods, the use of a vacuum-compatible DMA is a requirement as a process step for producing high-purity metallic particles. A vacuum-tight radial DMA (RDMA) has been developed and tested at low pressures. Its performance has been evaluated by using a commercial NANO-DMA as the reference. The performance of this low-pressure RDMA (LP-RDMA) in terms of the width of its transfer function is found to be comparable with that of other NANO-DMAs at atmospheric pressure and is almost independent of the pressure down to 30 mbar. It is shown that LP-RDMA can be used for the classification of nanometer-sized particles (5-20 nm) under low pressure condition (30 mbar) and has been successfully applied to nanoparticles produced by ablating FeNi at low pressures.

Automated cell viability assessment using a microfluidics based portable imaging flow analyzer

In this work, we report a system-level integration of portable microscopy and microfluidics for the realization of optofluidic imaging flow analyzer with a throughput of 450 cells/s. With the use of a cellphone augmented with off-the-shelf optical components and custom designed microfluidics, we demonstrate a portable optofluidic imaging flow analyzer. A multiple microfluidic channel geometry was employed to demonstrate the enhancement of throughput in the context of low frame-rate imaging systems. Using the cell-phone based digital imaging flow analyzer, we have imaged yeast cells present in a suspension. By digitally processing the recorded videos of the flow stream on the cellphone, we demonstrated an automated cell viability assessment of the yeast cell population. In addition, we also demonstrate the suitability of the system for blood cell counting. (C) 2015 AIP Publishing LLC.

Alvará, porque Vossa Alteza Real he servido revogar toda a prohibição, que havia, de fabricas, e manufacturas no Estado do Brazil, e dominios ultramarinos, na forma acima exposta : para Vossa Alteza Real João Alvares de Miranda Varejão o fez

Apresenta o alvará de 12 de abril de 1808, do Príncipe Regente D. João, que revoga o Alvará de 5 de janeiro de 1785, de Maria I, que extinguia a manufatura de tecidos no Brasil.

As obras do doctor Francisco de Saa de Miranda : agora de novo impressas com a relação de sua calidade, & vida, com todas as licenças necessarias por Por Vicente Aluarez : com privilegio real por dez annos

Francisco de Sá de Miranda nasceu provavelmente em Coimbra, em 1481e morreu em 1558. A posição de Sá de Miranda na poesia portuguesa é revolucionária, pelas novas formas que introduziu e pelos valores morais que defendeu. A ele são devidas as seguintes inovações: comédia em prosa, metro decassílabo, novas estruturas estróficas, novos subgêneros líricos. Quanto às intenções pedagógico-moralizantes há que assinalar, como fez Mourão Ferreira "a defesa dos valores castiços, a condenação dos novos tempos, a pregação de um regresso à natureza e a salvaguarda da liberdade moral". Sobre ele D. Carolina Michaelis já havia dito: "sem Miranda não tínhamos um Bernardes: sem Miranda não havia Ferreira; sem Miranda não florescia um Camões". Trata-se essa obra da segunda edição, póstuma como a primeira, de 1595. Durante muito tempo, a segunda edição foi subestimada, mas dela disse D. Carolina Michaelis de Vasconcelos: "é mais rica do que a outra, tem, além disso, a biografia de D. Gonçalo Coutinho, que é um subsídio precioso, e apresenta uma redação mais corrente, mais limada, uma fraseologia mais inteligível em várias partes, um sentido mais verossímil das passagens dificultosas". Acrescenta, ainda que "o lugar da impressão não se indica, mas é Lisboa". Das duas primeiras edições, disse Inocêncio: "qualquer delas é tida em conta de rara". Entretanto, esta edição tem sobre a primeira a vantagem de incluir uma biografia do autor atribuída por Barbosa Machado a D. Gonçalo Coutinho.

MBARI procedures for automated nutrient analyses using a modified Alpkem Series 300 Rapid Flow Analyzer

This report documents the methods used at the Monterey Bay Aquarium Research Institute (MBARI) for analyzing seawater nutrient samples with an Alpkem Series 300 Rapid Flow Analyzer (RFA) system. The methods have been optimized for the particular requirements of this laboratory. The RFA system has been used to analyze approximately 20,000 samples during the past two years. The methods have been optimized to run nutrient analyses in a routine manner with a detection limit of better than -±1% and a within run precision of -±1% of the full scale concentration range. The normal concentration ranges are 0-200 ~M silicate, 0-5 ~M phosphate, 0-50 ~M nitrate, 0-3 ~M nitrite, and 0-10 ~M ammonium. The memorandum is designed to be used in a loose-leaf binder format. Each page is dated and as revisions are made, they should be inserted into the binder. The revisions should be added into the binder. Retain the old versions in order to maintain a historical record of the procedures. (88 pages)

Alvará com força de Lei, pelo qual Vossa Alteza Real he servido crear hum Conselho Supremo Militar, e de Justiça, na forma acima declarada : para Vossa Alteza Real ver / João Alvares de Miranda Varejão o fez

Variação de Título: Criação do Conselho Supremo Militar

Los glacis de la cuenca de Miranda de Ebro (Burgos): significado climático y evolución geomorfológica

[ES] En este trabajo se analizan los glacis de la Cuenca de Miranda, en la que constituyen una de las morfologías de mayor relevancia paisajística. Se han establecido dos niveles de glacis cuyas características topográficas, morfológicas y sedimentológicas se integran plenamente dentro de los rasgos propios de estas morfologías. El significado ambiental de estas formaciones y su correlación con los depósitos aluviales del Ebro, han permitido establecer un marco cronológico para ambos niveles, asociando su génesis a los estadios isotópicos 2 y 4, y todo ello dentro de un marco regional coherente.

Caracterización de los depósitos fluviales del río Ebro en la cuenca de Miranda de Ebro (Burgos)

[ES] La evolución fluvial del río Ebro ha sido analizada de forma desigual, siendo más numerosas las investigaciones centradas en su curso medio y bajo. En este trabajo se describe el sistema de terrazas fluviales elaborado por el río Ebro a su paso por la Cuenca de Miranda de Ebro (Álava – Burgos). Se han reconocido cinco niveles de terrazas (T1-T5) mediante su análisis estratigráfico, morfológico y sedimentológico. Las terrazas se sitúan a unas alturas que oscilan entre los 68 y los 5 m sobre el nivel actual del río Ebro. El relleno sedimentario de los niveles más elevados (T1-T4) está compuesto por gravas poligénicas bien clasificadas y muy redondeadas con presencia puntual de niveles de arenas; mientras que el nivel más bajo (T5) está constituido enteramente por partículas finas. Los resultados apuntan hacia variaciones de caudal y aportes sedimentarios, en relación con las oscilaciones climáticas cuaternarias, como los factores que gobiernan la evolución fluvial en la Cuenca de Miranda de Ebro, en coherencia con los modelos de evolución propuestos para otros puntos de la cuenca hidrográfica

Universal Greenberger-Horne-Zeilinger-state analyzer based on two-photon polarization parity detection

We present a universal analyzer for the three-particle Greenberger-Horne-Zeilinger (GHZ) states with quantum nondemolition parity detectors and linear-optics elements. In our scheme, all of the three-photon GHZ states can be discriminated with nearly unity probability in the regime of weak nonlinearity feasible at the present state of the art experimentally. We also show that our scheme can be easily extended to the analysis of the multi-particle GHZ states.