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The effect of culture media and mycotoxin on porcine preimplantation embryonic development has been investigated. In the first experiment, porcine embryonic cleavage rates were similar in NCSU-23 and PZM-3 culture media, while more blastocysts were produced in PZM-3 (P<0.05). BAX and BCL2L1 transcription levels were similar in blastocysts cultured in both media. Cleavage rates were significantly decreased in the presence of cycloheximide (P<0.05), and both á-amanitin and cycloheximide completely inhibited blastocyst formation. In the mycotoxin experiment, porcine embryonic cleavage rates decreased in 10 ìM á-ZEA group, while blastocyst rates decreased in 30 ìM á-ZEA group (P<0.05). Total cell numbers of blastocysts were significantly lower in the 10 µM á-ZEA group (P<0.05). The transcription levels of POU5F1 and BCL2L1 were similar, while that of BAX and the ratio of BAX/BCL2L1 significantly increased in 3 ìM and 10 ìM á-ZEA groups compared to control group.

The diploma thesis deals with the use of meristem cultures in garlic (Allium sativum) breeding. The source material was used variety Tantalum of garlic. The using material, before the isolation of meristem, was tested for the occurrence of viral diseases by immunological tests ELISA (Enzyme-Linked Immuno-Sorbent Assay), also known as EIA (Enzyme Immunoassay). The method used to detect antibodies and antigens. The material was tested for viruses onion yellow dwarf (OYDV - Onion Yellow Dwarf Virus) and virus genus Carlavirus (GCLV - Common Garlic Latent Virus). Then the sound material was sterilized, cultured and then it was obtained meristem. Cultivation of preparation meristem was realised according available methodologies. After preparation meristem put on MS medium with vitamins and growth regulator IAA, NAA and BAP.

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Agradecemos a compreensão. on 2017-07-13T19:54:00Z (GMT) / Submitted by TAISE FLORÊNCIO null (taise_florencio@hotmail.com) on 2017-07-13T23:51:34Z No. of bitstreams: 1 Defesa - final.pdf: 1125308 bytes, checksum: 5b8d9f027a3241cc2a70f910280f3d13 (MD5) / Approved for entry into archive by Monique Sasaki (sayumi_sasaki@hotmail.com) on 2017-07-14T18:54:41Z (GMT) No. of bitstreams: 1 florencio_t_me_jabo.pdf: 1125308 bytes, checksum: 5b8d9f027a3241cc2a70f910280f3d13 (MD5) / Made available in DSpace on 2017-07-14T18:54:41Z (GMT). No. of bitstreams: 1 florencio_t_me_jabo.pdf: 1125308 bytes, checksum: 5b8d9f027a3241cc2a70f910280f3d13 (MD5) Previous issue date: 2017-06-23 / Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq) / Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP) / A biotecnologia utilizada no cultivo de microalgas à base de meio comercial é de alto custo. Sendo assim, alguns estudos indicam que o uso de meios de cultura alternativos com a finalidade de reduzir o preço de produção em curto espaço de tempo e mantendo alto valor nutricional, são procedimentos a serem adotados. O objetivo deste trabalho foi avaliar a qualidade de diferentes meios de cultivo a base de macrófitas. O meio com macrófita foi associado ao fertilizante inorgânico NPK (20:5:20). O experimento foi conduzido no período de 28 dias em triplicata. O meio de cultura com E. crassipes alcançou maior densidade celular (434 x 105 cel. mL-1), comparado com os outros meios de cultura com macrófitas que variaram de 319,7 x 105 cel. mL-1 (E. azurea) a 223,5 x 105 cel. mL-1 (T. domingensis). Durante o período experimental o oxigênio dissolvido, pH e condutividade elétrica não apresentaram diferenças significativas (P>0,05). A taxa de crescimento e densidade celular da microalga cultivada em meio de cultura com plantas aquáticas foram maiores (P>0,05) do que no meio NPK porém, similares (P<0,05) ao meio comercial CHU12. Em relação aos nutrientes, o teor de N (67 g.L-1) foi o mais elevado no meio de cultura com T. domingensis e os demais nutrientes (P, K, Ca, S, B, Cu, Fe e Zn) apresentaram concentrações abaixo de 35 g.L-1, nos diferentes meios utilizados. O uso de macrófitas como fonte alternativa para ser utilizada como meio de cultura no desenvolvimento de A. gracilis demonstrou ser viável tanto do ponto de vista econômico, nutricional e com elevada biomassa algal, sendo que E. crassipes, E. azurea e T. domingensis apresentaram os melhores resultados. / Since biotechnology in commercial medium-based culture of microalgae has very high costs, several studies recommend alternative medium cultures to reduce product costs in a short time period, coupled to high nutritional values. Objective of this work was to evaluate the quality of different macrophytes based culture media. Medium with macrophyte was associated with inorganic fertilizer NPK (20:5:20). Assay was conducted for 28 days, in triplicate. Culture medium with E. crassipes had the greatest cell density (434 x 105 cel. mL-1) when compared to other culture media with macrophytes and ranged between 319.7 x 105 cel. mL-1 (E. azurea) and 223.5 x 105 cel. mL-1 (T. domingensis). Dissolved oxygen, pH and electric conductivity did not show any significant differences during the experimental period (p>0.05). Growth rate and cell density of microalga cultivated in a medium with aquatic plants were greater (p>0.05) than in NPK medium, albeit similar (p<0.05) to the commercial medium CHU12. Further, N rate (67 g.L-1) was the highest in culture medium with T. domingensis, whilst the other nutrients (P, K, Ca, S, B, Cu, Fe and Zn) had lower than 35 g.L-1 concentrations in the different media employed. The use of macrophytes as an alternative source as culture medium in the development of A. gracilis proved to be viable from the economic and nutritional point of view, coupled to high algal biomass. E. crassipes, E. azurea and T. domingensis provided the best results. / CNPq: 130584/2015-0 / FAPESP: 2014/24697-3

The skeletal muscle is a dynamic tissue that has the ability to change and modify itself to fit the level of required activity; a phenomenon called muscle plasticity. Most studies of muscle plasticity are carried out in situ, a condition for which it is difficult to study and discern between the intrinsic properties of skeletal muscle, the myokines released by muscle fibers and the neurotrophic factors released by neurons innervating skeletal muscles that play various roles in the mechanisms of muscle plasticity. Another approach is to study the morphological and contractile properties of single adult muscle fibers under culture conditions for which one can fully control the level of activity and exogenous factors affecting muscle plasticity. However, the survival of single muscle fiber in culture is very low as most fibers degenerated or supercontracted within 5-7 days. The first objective of this study was to optimize fiber survival in culture. The application of chronic stimulation and beta-adrenergic agonists are two major factors that prevent muscle atrophy and loss of force in denervated muscles in situ. So, objective two was to determine if chronically stimulated single fibers in culture also improve fiber survival and contractile characteristic under culture conditions. The third objective was the same for salbutamol, a beta 2-adrenergic agonist. In regard to the optimization of fiber survival, the Minimum Essential Medium (MEM) was a better medium than Dulbecco’s Modified Eagle Medium (DMEM), changing 50% of the culture medium every two days also improved fiber survival compared to changing the medium every day. Interestingly, inhibiting the proliferation of satellite cells with AraC largely improved fiber survival when fibers were kept under resting conditions, but not when they were chronically stimulated. Finally, under conditions in which proliferation of satellite cells was inhibited, the use of a collagen/laminin mixture as adhering substrate to improve fiber adhesion to glass coverslip gave rise to a better fiber survival than Matrigel that contains not only collagen and laminin but several growth factors. The results suggest i) that when satellite cells (or fibroblasts) are allowed to proliferate they appear to contribute to the degeneration of fibers under resting conditions and ii) that the release of myokines by skeletal muscle fibers (or cytokines by other cells) likely play a role in fiber survival. Contrary to the situation in situ, neither the chronic stimulation nor salbutamol improved fiber survival and contractile characteristics of muscle fibers in culture suggesting that some important factors in culture are missing to allow chronic stimulation and salbutamol to reduce muscle atrophy and loss of force.

Statistically based experimental designs were applied to optimise medium constituent for biogas production utilizing N-­‐acetylglucosamine as a carbon source for Clostridium beijerinckii and Clostridium paraputrificum. The important medium constituents influencing total biogas produced, identified by the Plackett and Burman method, were FeSO4.7H2O and initial pH for C. beijerinckii cultures whilst for C. paraputrificum cultures N-­‐acetylglucosamine, L-­‐ cysteine.HCl.H2O and MgCl2. A one factor L-­‐cysteine.HCl.H2O optimization design was applied to investigate the ideal concentration of L-­‐cysteine.HCl.H2O required to achieve an anaerobic environment for optimum C. beijerinckii total biogas production. The Method of Steepest Ascent was then employed to locate the optimal area of the significant medium variables. Using the Box-­‐behnken method, experimental results showed that there were significant linear effects of independent variables, N-­‐acetylglucosamine for C. beijerinckii cultures and for C. paraputrificum cultures N-­‐acetylglucosamine, L-­‐cysteine.HCl.H2O and MgCl2 on total biogas volume. Significant curvature or quadratic effects of N-­‐ acetylglucosamine and L-­‐cysteine.HCl.H2O were identified for C. paraputrificum cultures. There were no significant interaction effects between medium constituent variables on resulting biogas volume. The optimal conditions for the maximum volume of biogas produced for C. beijerinckii cultures were 21 g/l of N-­‐ acetylglucosamine, 0.1 g/l of FeSO4.7H2O and initial pH of 6.11 and for C. paraputrificum were 29 g/l of N-­‐acetylglucosamine, 0.27 g/l of L-­‐ cysteine.HCl.H2O and 0.4 g/l of MgCl2. Using this statistical optimization strategy, the total biogas volume from N-­‐acetylglucosamine utilization increased from 150 ml/l to 6533 ml /l in the C. beijerinckii cultures and 100 ml/l to 5350 ml/l in the C. paraputificum cultures. The maximum yield of bio-­‐hydrogen by C. paraputrificum from N-­‐acetylglucosamine was 2.55 mol of H2 / mol of N-­‐ acetylglucosamine and by C. beijerinckii was 2.43 mol of H2 / mol of N-­‐ acetylglucosamine.

Epitélios confluentes de queratinócitos autólogos, cultivados segundo metodologia padronizada por Rheinwald e Green em 1975, têm sido usados como enxertos em diferentes situações clínicas. Contudo, a presença de componentes xenobióticos empregados nesta metodologia implica na possibilidade de transmissão de zoonoses, príons e viroses aos pacientes, além de envolver questões éticas relacionadas à utilização de animais. Tais preocupações têm direcionado pesquisadores a buscar alternativas que superem este impasse; no entanto, as formulações obtidas até o momento não são completamente satisfatórias. Assim, nossa proposta neste estudo, foi omitir ou substituir os componentes xenobióticos tradicionalmente utilizados no meio para queratinócitos, por similares humanos. Como resultado, padronizamos um meio de cultura onde omitimos o emprego de toxina colérica, substituímos o soro fetal bovino por lisado de plaquetas humanas na concentração de 2,5% e a insulina de origem bovina foi substituída por insulina recombinante humana na mesma concentração do método original (5 &mu;g/mL). Com os resultados obtidos foi possível concluir ser viável o cultivo de queratinócitos humanos, mantidos em meio de cultura livre de componentes xenobióticos. / Confluent epithelia of autologous keratinocytes, cultivated according to standardized methodology by Rheinwald and Green in 1975, have been used as grafts in different clinical situations. However, the presence of xenobiotics components applied in this method implies the possibility of transmission of zoonoses, príons, and viruses to patients, besides involving ethical issues related to the use of animals for components obtainment. Such concerns have driven researchers to seek alternatives that overcome this deadlock, as the formulations obtained so far are not completely satisfactory. Thus, our proposal in this study was to omit or replace the xenobiotics components traditionally used in the medium to keratinocytes culture, by human similar. As a result, we have standardized a culture medium whereby we omitted the use of cholera toxin, replaced fetal bovine serum by human platelet lysate at a 2.5% concentration and bovine insulin was replaced by recombinant human insulin at the same concentration as the original method (5 &mu;g/mL). With the results obtained we could conclude that the method to be viable to cultivate human keratinocytes, kept in culture medium free of xenobiotics components.

Epitélios confluentes de queratinócitos autólogos, cultivados segundo metodologia padronizada por Rheinwald e Green em 1975, têm sido usados como enxertos em diferentes situações clínicas. Contudo, a presença de componentes xenobióticos empregados nesta metodologia implica na possibilidade de transmissão de zoonoses, príons e viroses aos pacientes, além de envolver questões éticas relacionadas à utilização de animais. Tais preocupações têm direcionado pesquisadores a buscar alternativas que superem este impasse; no entanto, as formulações obtidas até o momento não são completamente satisfatórias. Assim, nossa proposta neste estudo, foi omitir ou substituir os componentes xenobióticos tradicionalmente utilizados no meio para queratinócitos, por similares humanos. Como resultado, padronizamos um meio de cultura onde omitimos o emprego de toxina colérica, substituímos o soro fetal bovino por lisado de plaquetas humanas na concentração de 2,5% e a insulina de origem bovina foi substituída por insulina recombinante humana na mesma concentração do método original (5 &mu;g/mL). Com os resultados obtidos foi possível concluir ser viável o cultivo de queratinócitos humanos, mantidos em meio de cultura livre de componentes xenobióticos. / Confluent epithelia of autologous keratinocytes, cultivated according to standardized methodology by Rheinwald and Green in 1975, have been used as grafts in different clinical situations. However, the presence of xenobiotics components applied in this method implies the possibility of transmission of zoonoses, príons, and viruses to patients, besides involving ethical issues related to the use of animals for components obtainment. Such concerns have driven researchers to seek alternatives that overcome this deadlock, as the formulations obtained so far are not completely satisfactory. Thus, our proposal in this study was to omit or replace the xenobiotics components traditionally used in the medium to keratinocytes culture, by human similar. As a result, we have standardized a culture medium whereby we omitted the use of cholera toxin, replaced fetal bovine serum by human platelet lysate at a 2.5% concentration and bovine insulin was replaced by recombinant human insulin at the same concentration as the original method (5 &mu;g/mL). With the results obtained we could conclude that the method to be viable to cultivate human keratinocytes, kept in culture medium free of xenobiotics components.

Orientador: Ricardo Alan Verdú Ramos / Resumo: A biomassa derivada de microalgas apresenta um grande potencial devido a sua sustentabilidade e alta produtividade, sendo possível extrair lipídios para produção de biodiesel. Entretanto, desafios na cadeia de produção como um todo devem ser resolvidos para que o biodiesel de microalgas seja viável. Uma das etapas críticas é o cultivo, sendo o meio de cultura um elemento de alto custo. O objetivo principal deste trabalho é avaliar o crescimento de microalgas visando produção de lipídios para biodiesel, utilizando como fontes de nutrientes a vinhaça originada do processo produtivo de etanol no setor sucroalcooleiro. Foram isolados e indentificados três gêneros nativos de microalgas: uma cianobactéria Aphanocapsa sp., uma clorofícea Oocystis sp. e outra clorofícea Scenedesmus sp. O cultivo da microalga Scenedesmus sp. em fotobiorreator de placas planas com meio de cultivo MBM (Modified Bristol Medium) se mostrou modesta em termos de produtividade de biomassa (8 mg/l.dia) e em teor de lipídios na biomassa seca (1,5%). O cultivo dessa mesma microalga em tubos de ensaios com meio alternativo utilizando vinhaça (três diluições de 2%, 5% e 10% em volume) no meio de cultura mostrou desempenho comparável em relação ao meio sintético MBM, sendo que a partir do dia 6, os quatros cultivos se estabilizam em torno de uma concentração celular de 6×106 de células/ml, indicando que a vinhaça pode ser uma fonte de nutrientes de baixo custo para o cultivo de microalgas. Deste modo, é possível r... (Resumo completo, clicar acesso eletrônico abaixo) / Mestre

Made available in DSpace on 2014-06-11T19:33:39Z (GMT). No. of bitstreams: 0 Previous issue date: 2010-05-27Bitstream added on 2014-06-13T20:25:30Z : No. of bitstreams: 1 sinimbuneto_fa_dr_jabo.pdf: 1254027 bytes, checksum: c24ee888986c7eaef05caa5e9f93c121 (MD5) / Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES) / O bacurizeiro (Platonia insignis Mart.) é uma frutífera nativa da Amazônia que apresenta alogamia acentuada e auto-incompatibilidade esporofítica. O objetivo desse trabalho foi estudar a germinação de pólens de bacuri por admitir-se tratar de uma técnica semelhante ao comportamento germinativo “in vivo”. A viabilidade “in vitro” tem sido utilizada para representar a capacidade potencial do pólen para completar o processo de fertilização. Os ensaios foram conduzidos em condições de laboratório em um delineamento inteiramente casualizado, analisados em esquema fatorial 2 x 3 x 4, onde: 2- formas de propagação (pé-franco e enxertada) X 3-estágios da antese (préantese, antese e pós-antese) X 4-concentrações de sacarose (0%; 7,5%; 10 e 20%), com 10 repetições. Para plantas de bacuri estudadas, há diferença de germinação do pólen, sendo que para polinização manual, o pólen deve ser coletado na antese, enquanto que o melhor meio para germinação é com 7,5 g 100 mL-1 de sacarose / The “bacurizeiro” (Platonia insignis Mart.) is an Amazon fruitful native tree that has marked allogamy and self-sporophytic incompatibility. The aim of this work was to study the pollen germination of “bacuri” to admit it is a similar technique to the germination behavior “in vivo”. The viability in vitro has been used to represent the potential ability of pollen to complete the fertilization process. The tests were conducted under laboratory conditions in a completely randomized design, tested in a 2 x 3 x 4 factorial, where: 2- plant forms (grafted and freestanding) X 3- pollen stage (pre-anthesis , anthesis and post anthesis) X-4 sucrose concentrations (0%, 7.5%, 10 and 20%), with 10 repetitions. For each bacurizeiro plant studied, there is difference in pollen germination, and for hand pollination, pollen should be collected at anthesis, while the best medium for germination is with 7,5 g 100 mL-1 of sucrose