Orientadores : Gil Eduardo Serra, Silvio Roberto Andrietta / Tese (doutorado) - Universidade Estadual de Campinas, Faculdade de Engenharia de Alimentos / Made available in DSpace on 2018-08-02T15:14:48Z (GMT). No. of bitstreams: 1 Pulzatto_MarciaEdilamar_D.pdf: 26357904 bytes, checksum: 2df0a217d125b100003208ac5fbc6c8b (MD5) Previous issue date: 2000 / Resumo: O resumo podera ser visualizado no texto completo da tese digital / Abstract: The abstract is available with the full electronic digital document / Doutorado / Doutor em Tecnologia de Alimentos

Orientadores: Valdemiro Carlos Sgarbieri, Vera Lucia Signoreli Baldini / Dissertação (mestrado) - Universidade Estadual de Campinas, Faculdade de Engenharia de Alimentos / Made available in DSpace on 2018-08-03T05:38:31Z (GMT). No. of bitstreams: 1 Rosa_LeonidiaLeite_M.pdf: 14450406 bytes, checksum: 66a23140212638978355d8ec62b59f35 (MD5) Previous issue date: 2003 / Resumo: O resumo poderá ser visualizado no texto completo da tese digital / Abstract: The abstract is available with the full electronic document / Mestrado / Mestre em Ciência da Nutrição

Orientador: Marcelo Cristianini / Dissertação (mestrado) - Universidade Estadual de Campinas, Faculdade de Engenharia de Alimentos / Made available in DSpace on 2018-08-03T20:55:51Z (GMT). No. of bitstreams: 1 Campos_FlavioPeckolt_M.PDF: 6100586 bytes, checksum: a6cb9cdaa3c3951640e77def65c8beab (MD5) Previous issue date: 2004 / Mestrado / Mestre em Tecnologia de Alimentos

No description available.

Orientador: Paulo Jose Samenho Moran / Dissertação (mestrado) - Universidade Estadual de Campinas, Instituto de Quimica / Made available in DSpace on 2018-08-03T16:09:51Z (GMT). No. of bitstreams: 1 Lourenco_Emerson_M.pdf: 1935914 bytes, checksum: 240e738140046c0d0084e84f77906305 (MD5) Previous issue date: 2003 / Mestrado

Orientadores: Glaucia M. Pastore, Gonçalo A. G. Pereira / Dissertação (mestrado) - Universidade Estadual de Campinas, Faculdade de Engenharia de Alimentos / Made available in DSpace on 2018-08-01T00:42:29Z (GMT). No. of bitstreams: 1 Prazeres_JanainaNicanuziados_M.pdf: 17689382 bytes, checksum: bf211097d639dcc5fdedaff55e93d768 (MD5) Previous issue date: 2002 / Resumo: Lipoxigenases (LOX) catalizam a oxigenação de ácidos graxos polinsaturados, como o ácido linoléico, em hidroperóxidos, os quais via a ação sequencial das hidroperóxido liases produzem os compostos voláteis de aroma responsáveis pelo aroma de frescor de frutas, como o aldeído C6, hexanal. Os cDNAs que codificam a lipoxigenase, isoenzima 1, e hidroperóxido liase foram expressados em Saccharomyces cerevisiae, através do vetor pYES 2 (construções pYES2-LOX-1 e pYES2-HPL). A análise da seqüência parcial de nucleotídeos do cDNA de LOX-1 mostrou 96-99 % de similaridade com LOX-1 de soja (G/ycine max), enquanto que a análise da seqüência completa de nucleotídeos do cDNA de HPL mostrou 97% de similaridade com HPL de pimentão (Capsicum annuum). A análise de Northem blot foi usada para caracterizar a expressão dos dois genes em S. cerevisiae duplo recombinates. A primeira análise pelo Northem blot mostrou que ambos os genes foram expressos após 4 horas de indução, contudo a segunda análise mostrou que somente um dos dois foram expressos nas leveduras duplo recombinantes, o que demonstrou uma baixa estabilidade dos vetores. A análise de cromatografia gasosa foi usada para medir as quantidades de hexanal produzido. A produção de hexanal foi limitada em todas as leveduras estudadas, inclusive nos controles. Isto pode estar relacionado à expressão de apenas um dos genes nas S. cerevisiae duplo recombinantes. / Abstract: Lipoxygenase (LOX) catalyze the oxigenation of polyinsaturated fatty acids, such as linoleic acid, into hydroperoxides, which, via the sequential action of hydroperoxide Iyases (HPL), produce the aroma volatile compounds responsible for the fresh fruit flavor, notably the C6 aldeyde, the hexanal. The cDNAs encoding soybean lipoxygenase isoenzyme 1 and hydroperoxide Iyase were expressed in Saccharomyces cerevisiae, using the pYES 2 vector (pYES2-LOX-1 and pYES2-HPL constructions). Analysis of partial nucleotide sequence of LOX-1 cDNA showed 96-99 % similarity with LOX-1 soybean (G/ycine max), while HPL cDNA full nucleotide sequence analysis showed 97 % similarity with HPL bell pepper (Capsicum annuum). Northern blot analysis was used to characterize the expression patterns for two genes in double recombinant S. cerevisiae. The first Northern blot analysis showed that both genes were expressed after 4 hours, however the second analysis showed that only one of them were expressed in the double recombinant yeast, what may suggest a low vector stability. Gas chromatography analysis was used to measure quantities of hexanal production, which was limited in ali yeasts studied, including the controls. Limited hexanal production can be related to the only one gene expression in the double recombinant S. cerevisiae. / Mestrado / Mestre em Ciência de Alimentos

No description available.

Seminario de Título entregado a la Universidad de Chile en cumplimiento parcial de los requisitos para optar al Título de Ingeniero en Biotecnología Molecular. / Los fotorreceptores se encuentran ampliamente distribuidos en todos los dominios de la vida, y se caracterizan por experimentar cambios de conformación inducidos por la presencia o ausencia de luz. Estas moléculas, junto con distintas herramientas ópticas, han permitido la aparición de la optogenética, que consiste en el uso de luz para permitir el comando de distintos procesos biológicos con un fino control espacio-temporal. El uso de luz como tratamiento inductor se destaca debido a que es una señal fácil de regular y con gran resolución espacial, popularizando su uso en sistemas mamíferos. Recientemente, el organismo modelo Saccharomyces cerevisiae ha sido también adoptado como una plataforma para este tipo de aproximaciones, ya que no presenta fotorreceptores descritos, y por lo tanto, no es capaz de sensar la luz. Recientemente, se ha descrito un sistema optogenético llamado Fungal Light- Oxygen-Voltage (FUN-LOV). Este se basa en la interacción de los dominios LOV de las proteínas WHITE COLLAR 1 (WC-1) y VIVID (VVD) del hongo Neurospora crassa, y en la arquitectura clásica de los sistemas de doble híbrido. FUN-LOV presentó notables niveles de inducción en la transcripción de un gen reportero luciferasa, y bajo ruido en condiciones de oscuridad, por lo que se presenta como uno de los sistemas optogenéticos más robustos reportados hasta la fecha. Sin embargo, poco se sabe sobre el papel que cumplen los distintos dominios de las proteínas que participan en este tipo de arquitectura para la funcionalidad y robustez del sistema optogenético. Es por esto que este seminario de título se presenta como una revisión de la modularidad y robustez del sistema FUN-LOV. Para esto se intercambiaron los dominios de unión a DNA (DBD) y de activación (AD) del sistema original por el de diferentes factores de transcripción (TF), ampliamente descritos en S. cerevisiae, tales como el DBD de las proteínas LexA y Cup2p, y el AD de VP16. Con el objetivo de realizar esta evaluación se generaron las correspondientes construcciones genéticas in silico, para luego ensamblarlas in vivo usando clonamiento por recombinación en levaduras. La evaluación de los sistemas se realizó de forma indirecta a través de la cuantificación de la actividad del gen reportero luciferasa (LUC) en respuesta a luz azul (BL) y al estímulo particular de cada dominio intercambiado (cobre, luz roja). Como resultado se obtuvo que ambos sistemas en los que se hizo un cambio a nivel de DBD/promotor, presentaron una pérdida en su funcionalidad y robustez, lo que sugiere fuertemente que dichos módulos son de vital importancia para este tipo de sistemas optogenéticos. Por su parte, el sistema FUN-LOV VP16 mantuvo su funcionalidad como interruptor-optogenético, pero presentó un menor nivel de inducción de actividad luciferasa, además de una cinética más lenta comparado con el sistema ya reportado. Finalmente, se concluye que el sistema FUN-LOV no es modular a nivel de DBD/promotor, ya que al remplazar estos módulos el sistema pierde su funcionalidad y robustez. Por otro lado, el sistema es modular a nivel de AD, ya que mantiene su funcionalidad, mientras que la robustez varía dependiendo de la naturaleza del AD a utilizar. / Photoreceptors are widely distributed in all the domains of life and are characterized by undergoing conformational changes induced by the presence or absence of light. These molecules, together with different optical tools, have allowed the appearance of optogenetics, which involves the use of light to allow the command of different biological processes with a fine spatio-temporal control. Light as an inducer treatment is remarkable since it is easy to regulate and provides great spatial resolution, which has boosted its use in mammalian systems. Recently, the model organism Saccharomyces cerevisiae has been adopted as an ideal platform for this kind of systems, due to the absence of described photoreceptors, and therefore, it is not capable to sense the light. Recently, an optogenetic system called FUN-LOV has been described. FUN-LOV is based on the interaction of Neurospora crassa photoreceptors WC-1 and VVD, and on the classical architecture of the double hybrid systems. FUN-LOV showed remarkable levels of luciferase gene expression, and low noise in dark conditions, so it is presented as one of the most robust optogenetic systems reported so far. Nonetheless, little is known about the role played by the different protein domains for the functionality and robustness of the optogenetic system. Therefore, this title seminar is presented as a revision of modularity and robustness of the FUN-LOV system. For this, the DBD and AD of the original system were exchanged for different TF domains widely described in S. cerevisiae, such as the DBD of the LexA and Cup2 proteins, and the AD of VP16. The evaluation was carried out generating the genetic constructions in silico, to then assemble them in vivo using yeast recombinational cloning. The evaluation of the systems was performed indirectly through the activity quantification of LUC reporter gene in response to BL and the particular stimulus for each domain exchanged (copper, red light). As results, a loss of functionality and robustness was observed when the domains where exchanged at the DBD/promoter level, strongly suggesting that these modules are crucial for this type of optogenetic systems. On other side, the FUN-LOV VP16 system keep its functionality as an opto-switch, but showed a lower induction of luciferase activity, and slower kinetics in relation with the already reported system. Finally, we concluded that FUN-LOV is not a modular system at the DBD/promoter level, because functionality and robustness of the system are lost when these modules were replaced. Nonetheless, the system is modular at the AD level, since it maintained its functionality, meanwhile the robustness varies depending on the nature of the AD used. / FONDECYT-Regular 1171151 y el Instituto Milenio de Biología Integrativa (iBio).

Heterochromatin in S. cerevisiae is formed at telomeres, rDNA, and the mating type loci by the Silent Information Regulator (SIR) complex. Silencing requires the SIR complex that consists of Sir2, Sir3, and Sir4. The SIR proteins interact with each other, nucleosomes, and DNA binding proteins that are located at silencers. Although the interactions within the SIR complex are well defined, the requirements for each of these interactions during the nucleation and spreading of heterochromatin are not. This study uses genetic and biochemical techniques to assess silencing at various loci and to detect interactions between the SIR proteins. Asf2 (Anti-Silencing Factor 2) is a poorly characterized protein that interacts with Sir3 and is investigated in detail throughout this work. The overexpression of ASF2 disrupts silencing and does so by outcompeting Sir4 for Sir3 binding. ASF2 is a paralog of SIR4, and they share significant homology within their coiled-coil domains which is required for their interaction with Sir3. The Asf2 protein exists as a dimer that depends on Sir3 and may serve as a tool to alter Sir3 availability and impact heterochromatin stability. The evidence presented here categorizes the requirements for the Sir3-Sir4 interaction and the establishment of H4K16 acetylation in nucleation and spreading. Mutations in the AAA+ domain of Sir3 (sir3-4A) render it insufficient to nucleate heterochromatin but do not prevent Sir3-4A and Sir4 from spreading downstream of silencers. The Sir3-Sir4 interaction is therefore a nucleation-specific requirement. Cells lacking SAS2 are defective for telomere silencing, but silencing is partially restored by overexpressing SIR3 but not sir3-4A. Although the Sir3-Sir4 interaction is not required for Sir3 to spread on its own, Sir4/Sir2 are unable to spread without the establishment of H4K16 acetylation.

No description available.