Producción in vitro de Embriones Bovinos Transgénicos mediante Inyección Intracitoplasmática de Espermatozoides (ICSI) con Enzimas de Restricción. In vitro Production of Transgenic Bovine Embryos using Intracitoplasmic Sperm Injection (ICSI) and Restriction Enzymes

La ingeniería genética y la reprogramación de organismos vivos representan las nuevas fronteras biotecnológicas que permitirán generar animales con modificaciones precisas en sus genomas para un sinnúmero de aplicaciones biomédicas y agropecuarias. Las técnicas para inducir modificaciones génicas intencionales en animales, especialmente en especies mayores de interés agropecuario, se encuentran rezagadas si se compara con los avances significativos que se han producido en el área de la transgénesis de roedores de laboratorio, especialmente el ratón. Es así que, el presente proyecto persigue desarrollar y optimizar protocolos para generar embriones bovinos transgénicos para aplicaciones biotecnológicas. La estrategia propuesta, se basa en conseguir la presencia simultánea en el interior celular de una enzima de restricción (I-SceI) más un transgén (formado por casetes de expresión de una proteína fluorescente -ZsGreen1- y neomicina fosfotransferasa). Específicamente, proyectamos estudiar una vía alternativa para generar embriones bovinos transgénicos mediante la incorporación del transgén (casetes ZsGreen1 y neo) flanqueado por sitios I-SceI más la enzima I-SceI al interior del ovocito junto con el espermatozoide durante la técnica conocida como inyección intracitoplasmática de espermatozoides (ICSI). Los embriones así generados se cultivarán in vitro, inspeccionándolos diariamente para detectar la emisión de fluorescencia, indicativa de la expresión de la proteína ZsGreen1. Los embriones que alcancen el estado de blastocisto y expresen el transgén se transferirán quirúrgicamente al útero de ovejas sincronizadas y se mantendrán durante 7 días. Al cabo de este período, los embriones se recolectarán quirúrgicamente del útero ovino y se transportarán al laboratorio para determinar el número de sitios de integración y número de copias del transgén mediante el análisis de su ADN por Southern blot. Se prevé que los resultados de esta investigación permitirán sentar las bases para el desarrollo de métodos eficientes para obtener modificaciones precisas en el genoma de los animales domésticos para futuras aplicaciones biotecnológicas. Genetic engineering and reprogrammed organisms represent the new biotechnological frontiers which will make possible to generate animals with precise genetic modifications for agricultural and biomedical applications. Current methods used to generate genetically modified large animals, lay behind those used in laboratory animals, specially the mouse. Therefore, we seek to develop and optimize protocols to produce transgenic bovine embryos through the use of a non-viral vector. The strategy involves the simultaneous presence inside the cell of a restriction enzyme (I-SceI) and a transgene (carrying cassettes for a fluorescent protein -ZsGreen1- and neomycin phosphotransferase) flanked by restriction sites for the endonuclease. We plan to develop an alternative approach to generate transgenic bovine embryos by coinjecting the transgene flanked by I-SceI restriction sites plus the enzyme I-SceI along with the spermatozoon during the technique known as intracytoplasmic sperm injection (ICSI). Embryos will be cultured in vitro and inspected daily with a fluorescence microscope to characterize transgene expression. Embryos that reach the blastocyst stage and express the transgene will be surgically transfer to the uterus of a synchronized ewe. After 7 days, the embryos will be flushed out the ovine uterus and transported to the laboratory to determine the number of integration sites and transgene copies by Southern blot. We anticipate that results from this research will set the stage for the development of efficient strategies to achieve precise genetic modifications in large domestic animals for future biotechnological applications.

Prevalência de Fatores de Risco Cardiovascular em Pacientes em Hemodiálise - O Estudo CORDIAL

Fundamentos: Há uma carência de dados epidemiológicos sobre o perfil de risco cardiovascular nos pacientes renais crônicos em hemodiálise no Brasil. Objetivo: O estudo CORDIAL foi planejado para avaliar fatores de risco cardiovascular e acompanhar a evolução de uma população em programa de hemodiálise numa cidade metropolitana do Brasil. Métodos: Todos os pacientes em hemodiálise por doença renal crônica nos quinze centros de nefrologia de Porto Alegre foram considerados para inclusão na fase inicial do estudo CORDIAL. Dados clínicos, laboratoriais e demográficos foram obtidos nos registros médicos, e em entrevistas individuais estruturadas realizadas com todos os pacientes por pesquisadores treinados. Resultados: Foram incluídos 1215 pacientes (97,3% de todos os que estavam em hemodiálise na cidade de Porto Alegre). A média de idade era 58,3 anos, 59,5% eram homens e 62,8% eram brancos. A prevalência de fatores de risco cardiovascular encontrada foi 87,5% para hipertensão, 84,7% para dislipidemia, 73,1% para sedentarismo, 53,7% para tabagismo e 35,8% para diabetes. Em uma análise multivariada ajustada, sedentarismo (p = 0,032; RP 1,08 - IC95%: 1,01-1,15), dislipidemia (p = 0,019; RP 1,08 - IC95%: 1,01-1,14), e obesidade (p < 0,001; RP 1,96 - IC95%: 1,45-2,63) foram mais frequentes em mulheres; e hipertensão (p = 0,018; PR 1,06 - IC95%: 1,01-1,11) e tabagismo (p = 0,006; RP 2,7 - IC95%: 1,79-4,17) foram mais frequentes naqueles com menos de 65 anos. Sedentarismo apresentou uma associação independente com tempo em diálise inferior a 12 meses (p < 0,001; RP 1,23 - IC95%: 1,14-1,33). Conclusão: Pacientes em hemodiálise nesta metrópole do sul do Brasil apresentaram uma prevalência elevada de fatores de risco cardiovascular similar a diversos países do hemisfério norte.

Impuls und Transfer : literarische Feldbildungsprozesse in Magdeburg im 17. und 18. Jahrhundert

Magdeburg, Univ., Fak. für Geistes-, Sozial- und Erziehungswiss., Diss., 2007

Efeitos do arsênico sôbre a cultura do algodoeiro em terra arenosa

The authors studied the action of arsenic, in the form of lead arsenate and sodium arsenite, on cotton in white sandy soil of Piracicaba, State of S. Paulo, Brazil. The experiment was carried out in Mitscherlich pots, applying increasing quantities of the above mentioned compounds. The following conclusions were reached: sodium arsenite is more toxic than lead arsenate. 48 pounds per acre of lead arsenate and 16 pounds per acre of sodium arsenite reduced the vegetative development and the production of cotton. The roots were more seriously affected than the aerial parts. Sandy soils were sensitive to arsenic toxicity. The arsenic mobilization in the soil seems to depend upon factors such as, the a- cidity, the concentration of Fe2O3, CaO, P2O5 and soil colloids, both clay and humus components. The authors suggest, based on their own experiment and after a detailed study of the literature, the use of organic insecticids which may not leave toxic residues, rotation of crops, application of lime and reduction of arsenical sprays to a mini mum. Arsenic compounds should not be used in soils destined to the cultivation of food plants. Rice should not be planted in soils contaminated by arsenic compounds during several years of cotton cultivation. Future experiments are planed, using other soils such as "terra roxa", in Mitscherlich pots and in field plots.

Microdeterminação do arsênico

Studying the application of those methods of microanalysis which avoid costly instalations and atempting to combine high precision with low cost, the author recomends a new method consisting of the following : a) exposure of a surface of 530.66 mm2 of Zn to the action of the acid. b) instalation of 3 series of HgBr2 paper in test tubes with an internal diameter of respectively 3,5 and 9 mm. c - mouting between two slides, covering the margins (with parafin etc.) with parafin in order to conserve the results of the determination without change due to the action of light or moisture. d) the results can be compared at a level of 0.00001 mgr. A203 or 0.000007575 mgr. As.

Arsenicais e inseticidas orgânicos sôbre o algodoeiro em terras roxa e arenosa

The authors carried out a series of pots and plots experiments applying arsenical and organic insecticides to cotton plants cultivated in "terra roxa" and in a sandy soil. The first results were presented in 1947, to the la. Reunião Brasileira de Ciência do Solo (First Brazilian Congress of Soil Science); they pointed out the danger resulting from the accumulation of arsenic in soils due to the constant applications of arsenicais to control cotton pests; in the course of the time, the amount of residual arsenic in the soil would determine a decrease in cotton yield caused by its toxic effect on the crop. The following conclusions were drawn from the last three experiments: 1) the field experiment conducted in a sandy soil to which lead arseniate was applied in increasing rates produced a reduction of 50 per cent in the yield (the three highest doses were responsible for this result); by this way, the pot experiment published in 1947 was confirmed); 2) in the pot experiment with "terra roxa" toxic effects appeared only in the plants receiving the last dosis of lead arsenate; this result is explained quite naturally by a considerable absorption of the AsO4 --- ion by "terra roxa" colloidal material; furthermore the CaO, P2O5 and Fe2O3 content and the pH value (higher) would decrease the arsenate solubilization in the soil considered; 3) the pot experiment with organic insecticides applied in the rates usually employed in the control of cotton pests, showed that 10% D.D.TD. and 2.5% Rotenone did not affect cotton plants cultivated in a sandy soil; however we agree with FOSTER (1951), in the point that both mineral and organic insecticides must be applied in the minimum amount as possible; we also think that experiments like those should be carried out with the known insecticides, in several soil conditions and with many crops in order to determine the maximum limits of tolerancy.

Entwicklungsprojekt MCUI13 : Machbarkeitsstudie/ Konzept zum Bau und Betrieb einer „Power to Methanol“-Anlage

[s.c.]

Befristete Arbeitsverträge : Ein praxisorientierter Leitfaden

This master thesis explains fixed-term contracts in practice. To illustrate this point, examples are provided for better understanding. It will be discussed at both the fixed-term with an objective reason and without objective reason. The second variant is also called moderate Expiration Calendar. Not only the benefits and advantages for the employer are enumerated, but also the special challenges and problems that may arise. Particularly with regard to the prohibition of previous employment.This thesis presents the interests fo employers and companies. The central research question is: What are the possibilities of limitation and there is the prohibition of previous employment unconstitutional? A special attention is given to the contractual formulation examples of limited contracts. It deals exclusively with applicable German and European law. A comparison with previous case-law does not take place in this master thesis.

Doses and methods of distribution of potassium chloride on soybean crop (Glycine max (L.) Merrill) in a red latosol, affecting soil salinity, grain production and chemical composition of leaves

The present work deal t wi th an experiment under field conditions and a laboratory test of soil incubation the objectives were as follows: a. to study effects on soybean grain product ion and leaf composition of increasing doses of potassium chloride applied into the soil through two methods of distribution; b. to observe chemical modifications in the soils incubated with increasing doses of potassium chloride; and, c. to correlate field effects with chemical alterations observed in the incubation test, The field experiment was carried out in a Red Latosol (Haplustox) with soybean cultivar UFV - 1. Potassium chloride was distributed through two methods: banded (5 cm below and 5 cm aside of the seed line) and broadcasted and plowed-down. Doses used were: 0; 50; 100 and 200 kg/ha of K2O. Foliar samples were taken at flowering stage. Incubation test were made in plastic bags with 2 kg of air dried fine soil, taken from the arable layer of the field experiment, with the following doses of KC1 p,a. : 0; 50; 100; 200; 400; 800; 1,600; 3.200; 6,400 and 12,800 kg/ha of K(2)0. In the conditions observed during the present work, results allowed the following conclusions: A response by soybean grain production for doses of potassium chloride, applied in both ways, banded or broadcasted, was not observed. Leaf analysis did not show treatment influence over the leaf contents for N, P, K, Ca, Mg, and CI, Potassium chloride salinity effects in both methods of distribution for all the tested closes were not observed.

Action of growth regulators on soybean cultivar Davis

The effects of growth regularots on soybean plant (Glycine max) under greenhouse conditions were studied. Before flower ing, Agrostemmin (1 g/10 ml/3 1), gibberellic acid (GA) 100 ppm, and (2-chloroethyl) trimethylammonium chloride (CCC) 2,000 ppm were applied. At the flower anthesis, 2,3,5 - triio dobenzoic acid (TIBA) 20 ppm was applied. Other two applications with TIBA, with intervals of four days, were realized. Treatment with GA increased plant height while CCC presented a tendency to reduce it. Numbers of leaves, internods, and stems were not affected by the growth regulators.