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  • About
  • The Global ETD Search service is a free service for researchers to find electronic theses and dissertations. This service is provided by the Networked Digital Library of Theses and Dissertations.
    Our metadata is collected from universities around the world. If you manage a university/consortium/country archive and want to be added, details can be found on the NDLTD website.
91

Investigação dos determinantes moleculares envolvidos na interação com os substratos de Tsa1 e Tsa2 de Saccharomyces cerevisiae

Breyer, Carlos Alexandre 25 November 2015 (has links)
Submitted by Luciana Sebin (lusebin@ufscar.br) on 2016-09-20T18:14:03Z No. of bitstreams: 1 TeseCAB.pdf: 11456750 bytes, checksum: 7ab706a3e4cc604844c73a7452eece9a (MD5) / Approved for entry into archive by Marina Freitas (marinapf@ufscar.br) on 2016-09-21T12:48:53Z (GMT) No. of bitstreams: 1 TeseCAB.pdf: 11456750 bytes, checksum: 7ab706a3e4cc604844c73a7452eece9a (MD5) / Approved for entry into archive by Marina Freitas (marinapf@ufscar.br) on 2016-09-21T12:49:01Z (GMT) No. of bitstreams: 1 TeseCAB.pdf: 11456750 bytes, checksum: 7ab706a3e4cc604844c73a7452eece9a (MD5) / Made available in DSpace on 2016-09-21T12:49:11Z (GMT). No. of bitstreams: 1 TeseCAB.pdf: 11456750 bytes, checksum: 7ab706a3e4cc604844c73a7452eece9a (MD5) Previous issue date: 2015-11-25 / Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP) / Peroxiredoxins (Prx) are antioxidant proteins capable of decomposing a variety of hydroperoxide are very abundant in the cell and in eukaryotes are distributed in many cell compartments. The Prx are able to reduce their substrates using a highly reactive cysteine residue, named peroxidatic cysteine (CP-S-), which is maintained in the thiolate form as a consequence of the active site environment. In hydroperoxide decomposition process the CP is oxidized to cysteine sulfenic acid (CP-SOH). These vast majorities of these proteins are obligate dimers and has a second cysteine involved in the catalytic cycle, named resolution cysteine (CR), which forms a disulfide with CP and is often reduced by the enzyme thioredoxin (Trx). In addition to the reduction of Prx, the Trx are involved in several other biological processes such as cell growth, inhibition of apoptosis, transcriptional activation and DNA synthesis. When cell are exposed to oxidative stress, the CP can suffer overoxidation, forming species such as cysteine sulfinic acid (CP-SO2H) and cysteine sulfonic acid (CP-SO2H), which are not reduced by Trx. However, the CP overoxidation results in a change of the of Prx quaternary structure, resulting in the formation of high molecular weight structures (HMW) with molecular chaperone property and related to signal transduction triggered by hydroperoxides. Saccharomyces cerevisiae, have two isoforms of cytosolic Prx (Tsa1 and Tsa2), which shares high similarity (86% identity and 96% similarity), but differ in cellular abundance and expression levels. However, these enzymes are often considered redundant proteins. This study aimed the comparative analysis searching for functional and structural differences between Tsa1 and Tsa2. We evaluated Tsa1 and Tsa2 relationships with its oxidizing (hydroperoxide) and reducing substrates (Trx1 and Trx2). We also performed structural and kinetic analysis showed a significant relationship between maintaining the decameric structure the activity of enzymes which occurs by a series polar interactions but not equal to the active site amino acids. Highlighting the Thr44 of Tsa1 (Ser in Tsa2) which is involved in decamer structure stabilization by a CH-π interaction type with the Tyr77 and through the Oγ atom (from Thr) polar interactions with Phe45 and Leu41. Overoxidation susceptibility assays were performed using Tsa1 and Tsa2, and our results revealed that organic hydroperoxides were able to promote the CP overoxidation more efficiently when compared to H2O2. In turn, Tsa2 was very more resistant to CP overoxidation than Tsa1. This difference was also investigated concerning the catalytic triad Thr (Tsa1) to Ser (Tsa2). We generated mutants carrying reciprocal substitutions Tsa1T44S and Tsa2S44T and our results revealed that Tsa1T44S become more resistant to CP overoxidation and Tsa2S44T become more sensitive. The formation of high molecular weight structures (HMW) of Tsa1, Tsa2 and mutants were also investigated by size exclusion chromatography (SEC) and transmission electron microscopy (TEM). The Tsa1, Tsa2 and mutants HMW formation were analyzed by size exclusion chromatograph (SEC) and transmission electron microscopy (TEM). The results showed an increase of HMW formation after Trx system reaction using high concentrations of CHP. Were observed the stacking of the ring-shaped structures besides spherical species. Our results demonstrate that Tsa1 and Tsa2 proteins differ significantly in overoxidation susceptibility and HMW complex formation, indicating that these proteins have not redundant biological roles and small changes in active site promote high functional and structural changes in these enzymes. / Peroxirredoxinas (Prx) são proteínas antioxidantes capazes de decompor uma grande variedade de hidroperóxidos, são muito abundantes na célula e em eucariotos estão distribuídas nos diversos compartimentos celulares. As Prx são capazes de reduzir seus substratos utilizando um resíduo de cisteína altamente reativa, denominada de cisteína peroxidásica (CP-S-), que se apresenta na forma de tiolato. No processo de decomposição de hidroperóxidos é oxidada a cisteína ácido sulfênico (CP-SOH). A grande maioria destas proteínas se apresenta como dímeros obrigatórios e apesar de algumas Prx apresentarem somente uma cisteína, grande parcela possui uma segunda cisteína envolvida no ciclo catalítico que recebe o nome de cisteína de resolução (CR), a qual forma um dissulfeto com CP, que frequentemente é reduzido pela enzima tiorredoxina (Trx). Adicionalmente à redução das Prx, as Trx estão envolvidas em diversos outros processos biológicos como crescimento celular, inibição de apoptose, ativação de transcrição e síntese de DNA. Quando a célula é exposta a elevado estresse oxidativo, pode ocorrer a superoxidação de CP formando espécies superoxidadas como a cisteína ácido sulfínico (CP-SO2H) e sulfônico (CPSO3H) que não podem ser reduzidas por Trx. Entretanto, também resulta na alteração da estrutura quaternária das Prx, levando à formação de estruturas de alto peso molecular (HMW) que possuem propriedade de chaperona molecular e estão relacionadas a transdução de sinal desencadeada por hidroperóxidos. Em Saccharomyces cerevisiae, há duas isoformas citosólicas de Prx (Tsa1 e Tsa2), que possuem grande semelhança (86% de identidade e 96% de similaridade), mas apresentam diferenças na concentração celular e nível de expressão. Entretanto, muitas vezes são consideradas proteínas redundantes. Este trabalho teve como objetivos uma análise comparativa aprofundada buscando um maior entendimento das diferenças funcionais e estruturais de Tsa1 e Tsa2. Foram avaliados a relação de Tsa1 e Tsa2 com seus substratos oxidantes (hidroperóxidos) e substratos redutores (Trx1 e Trx2). Análises estruturais e cinéticas demonstraram uma importante relação entre a manutenção da estrutura decamérica a atividade das enzimas a qual ocorre por uma serie interações polares similares mas não iguais entre aminoácidos do sítio ativo. Com destaque para a Thr44 de Tsa1 (Ser em Tsa2) que possui importância na estabilização da estrutura decamérica através de uma interação do tipo C-H-π com Tyr77 e através do átomo de O que possui interações polares com Phe45 e Leu41. Também foram realizadas análises de susceptibilidade a superoxidação de Tsa1 e Tsa2, e demonstrou-se que peróxidos orgânicos são capazes de promover mais eficientemente a superoxidação quando comparados a H2O2, sendo que Tsa1 apresenta uma maior sensibilidade a superoxidação. Essa diferença foi também relacionada à substituição Thr/Ser e análises de susceptibilidade a superoxidação dos mutantes Tsa1T44S e Tsa2S44T demonstraram que a substituição reciproca tornou Tsa1T44S mais resistente a superoxidação e Tsa2S44T mais sensível. A formação de estruturas de alto peso molecular (HMW) de Tsa1, Tsa2 e mutantes foram investigadas através de cromatografia de exclusão molecular (SEC) e microscopia eletrônica de transmissão (TEM). Os resultados demonstraram um aumento da formação de HMW após reações de superoxidação utilizando o sistema Trx em altas concentrações de CHP e foi verificada a presença de empilhamentos de decâmeros, além de esferas, descritas na literatura. Os resultados obtidos neste trabalho demonstram de forma clara que as proteínas Tsa1 e Tsa2 diferem de forma significativa tanto na suscetibilidade a superoxidação quanto na formação de complexos HMW distintos, indicando fortemente que estas proteínas possuem papeis biológicos não redundantes e que alterações sutis de aminoácidos no sitio ativo promovem grandes alterações funcionais e estruturais nas enzimas.
92

Expressao, purificacao e caracterizacao de tireotrofina recombinante humana (rec-hTSH) em celulas de ovario de hamster chines (CHO)

MENDONCA, FERNANDA de 09 October 2014 (has links)
Made available in DSpace on 2014-10-09T12:48:54Z (GMT). No. of bitstreams: 0 / Made available in DSpace on 2014-10-09T14:01:11Z (GMT). No. of bitstreams: 1 09617.pdf: 5595560 bytes, checksum: 5a054c653d88ec87b2d3c979b79b00b5 (MD5) / Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP) / Dissertacao [Mestrado] / IPEN/D / Instituto de Pesquisas Energeticas e Nucleares - IPEN/CNEN-SP / FAPESP:00/09008-4
93

Estudo da polimerizacao do acetato de vinila utilizando a radiacao ionizante

MESQUITA, ANDREA C. 09 October 2014 (has links)
Made available in DSpace on 2014-10-09T12:47:50Z (GMT). No. of bitstreams: 0 / Made available in DSpace on 2014-10-09T14:08:26Z (GMT). No. of bitstreams: 1 08511.pdf: 4385457 bytes, checksum: a0c4d3d31c582b2a850f8616fec84e9e (MD5) / Dissertacao (Mestrado) / IPEN/D / Instituto de Pesquisas Energeticas e Nucleares - IPEN/CNEN-SP
94

Understanding Viscoelastic Behavior of Asphalt Binders Through Molecular Structure Investigation

January 2018 (has links)
abstract: Asphalt binder is a complex viscoelastic hydrocarbon, whose performance depends upon interaction between its physical and chemical properties, both of which are equally important to the successful understanding of the material. Researchers have proposed various models linking linear viscoelastic (LVE) and microstructural parameters. However, none of these parameters provide insight into the relationship in the non- linear viscoelastic NLVE domain. The main goals of this dissertation are two fold. The first goal is to utilize the technique of Laser Desorption Mass Spectroscopy (LDMS) to relate the molecular structure of asphalt binders to its viscoelastic properties. The second goal of the study is to utilize different NLVE characterization tools and analysis procedures to get a clear understanding of the NLVE behavior of the asphalt binders. The goals of the study are divided into four objectives; 1) Performing the LDMS test on asphalt binder to develop at the molecular weight distributions for different asphalts, 2) Characterizing LVE properties of Arizona asphalt binders, 3) Development of relationship between molecular structure and linear viscoelasticity, 4) Understanding NLVE behavior of asphalt binders through three different characterization methods and analysis techniques. In this research effort, a promising physico-chemical relationship is developed between number average molecular weight and width of relaxation spectrum by utilizing the data from LVE characterization and the molecular weight distribution from LDMS. The relationship states that as the molecular weight of asphalt binders increase, they require more time to relax the developed stresses. Also, NLVE characterization was carried out at intermediate and high temperatures using three different tests, time sweep fatigue test, repeated stress/strain sweep test and Multiple Stress Creep and Recovery (MSCR) test. For the intermediate temperature fatigue tests, damage characterization was conducted by applying the S-VECD model and it was found that aged binders possess greater fatigue resistance than unaged binders. Using the high temperature LAOS tests, distortion was observed in the stress-strain relationships and the data was analyzed using a Fourier transform based tool called MITlaos, which deconvolves stress strain data into harmonic constituents and aids in identification of non-linearity by detecting higher order harmonics. Using the peak intensities observed at higher harmonic orders, non-linearity was quantified through a parameter termed as “Q”, which in future applications can be used to relate to asphalt chemical parameters. Finally, the last NLVE characterization carried out was the MSCR test, where the focus was on the scrutiny of the Jnrdiff parameter. It was found that Jnrdiff is not a capable parameter to represent the stress-sensitivity of asphalt binders. The developed alternative parameter Jnrslope does a better job of not only being a representative parameter of stress sensitivity but also for temperature sensitivity. / Dissertation/Thesis / Doctoral Dissertation Civil, Environmental and Sustainable Engineering 2018
95

Determinação de espectros de relaxação e distribuição de massa molar de polímeros lineares por reometria

Farias, Thais Machado January 2009 (has links)
A distribuição de massa molar (DMM) e seus parâmetros são de fundamental importância na caracterização dos polímeros. Por este motivo, o desenvolvimento de técnicas que permitam a determinação da DMM de forma mais rápida e a menor custo é de grande importância prática. Os principais objetivos deste trabalho foram a implementação de alguns dos modelos baseados da teoria da reptação dupla propostos na literatura para descrever o mecanismo de relaxação das cadeias poliméricas, a avaliação dessas implementações e a análise de dois passos fundamentais na obtenção da DMM a partir de dados reológicos que são a metodologia de cálculo do espectro de relaxação baseado no modelo de Maxwell e a estratégia para a avaliação numérica das integrais que aparecem nos modelos de relaxação. Foi resolvido o problema denominado problema inverso, ou seja, a determinação da DMM a partir de dados reológicos usando um modelo de relaxação especificado e uma função de distribuição imposta. Foi usada a função Exponencial Generalizada (GEX) para representar a probabilidade de distribuição, sendo consideradas duas abordagens: i) cálculo explícito do espectro de relaxação e ii) aproximações paramétricas de Schwarzl, que evitam a necessidade do cálculo explícito do espectro de relaxação. A metodologia de determinação da DMM foi aplicada para amostras de polietileno e foram estimadas distribuições com boa representação dos dados experimentais do GPC, ao considerarem-se amostras com polidispersões inferiores a 10. Com relação a metodologia de cálculo do espectro de relaxação, foi realizado um estudo comparativo da aplicação de espectros de relaxação discreto e contínuo, com o objetivo de estabelecer critérios para especificação do número ótimo de modos de Maxwell a serem considerados. Ao efetuar-se a comparação entre as técnicas, verificou-se o espectro discreto apresenta como um sistema melhor condicionado, permitindo assim obter maior confiabilidade dos parâmetros estimados. Também é proposta uma modificação da metodologia de determinação da DMM, em que é aplicada a quadratura de Gauss-Hermite para a resolução numérica da integral dos modelos de relaxação. / The molecular weight distribution (MWD) and its parameters are of the fundamental importance in the characterization of polymers. Therefore, the development of techniques for faster and less time consuming determination of the MWD is of great practical relevance. The goals of this work were the implementation of some of the relaxation models from double reptation theory proposed in the literature, the evaluation of these implementations and the analysis of two key points in the recovery of the MWD from rheological data which are the methodology for calculation of the relaxation spectrum based on the Maxwell model and the numeric strategy for the evaluation of the integrals appearing in the relaxation models. The inverse problem, i.e., the determination of the MWD from rheological data using a specified relaxation model and an imposed distribution function, was solved. In the analysis of the inverse problem, the Generalized Exponential (GEX) was used as distribution function and two approaches were considered: i) explicit calculation of the relaxation spectrum and ii) use of the parametric method proposed by Schwarzl to avoid the explicit calculation of the relaxation spectrum. In the test of commercial samples of polyethylene with polidispersity less than 10, the application of this methodology led to MWD curves which provided good fit of the experimental SEC data. Regarding the methodology for calculation of the relaxation spectrum, a comparison between the performance of discrete and continuous relaxation spectrum was performed and some possible a criteria to determine the appropriate number of relaxation modes of Maxwell to be used were evaluated. It was found that the technique of discrete spectrum leads to better conditioned systems and, consequently, greater reliability of the estimated parameters. With relation to the numeric strategy for the evaluation of the integrals appearing in the relaxation models, the use of Gauss-Hermite quadrature using a new change of variables was proposed.
96

Produção de acido hialuronico por Streptococcus : estudo da fermentação e caracterização do produto / Production of hyaluronic acid by Streptococcus : study of the fermentation and characterization of the product

Ogrodowski, Christiane Saraiva 22 February 2006 (has links)
Orientador: Maria Helena Andrade Santana / Tese (doutorado) - Universidade Estadual de Campinas, Faculdade de Engenharia Quimica / Made available in DSpace on 2018-08-06T05:31:54Z (GMT). No. of bitstreams: 1 Ogrodowski_ChristianeSaraiva_D.pdf: 2070554 bytes, checksum: 434797b23387d845e6ccd6232e5a2e99 (MD5) Previous issue date: 2006 / Resumo: Neste trabalho estudou-se a produção de ácido hialurônico (AH) por via microbiana, correlacionando-se as variáveis do processo com a produtividade e propriedades do produto. Foram utilizas cepas de Streptococcus em fermentações descontínuas, e buscou-se obter o AH de massa molar elevada, da ordem de milhões de Da, com alta viscosidade e viscoelasticidade. Essas propriedades são requeridas para as aplicações médicas do AH no tratamento de doenças degenerativas e inflamatórias das articulações dos ossos, na reposição do fluido sinovial e como veículo de liberação de fármacos em implantes cirúrgicos. Foram avaliadas três cepas de mutantes de Streptococcus (S. equi, ATCC 39506 e 6580, e S. zooepidemicus ATCC 39920), crescidas em meio de cultura composto de glicose, extrato de leveduras e sais. A maior produção do AH com alta viscosidade e viscoelasticidade foi obtida com a cepa ATCC 39920. O planejamento fatorial completo de experimentos 22, foi utilizado para otimização do meio de cultura e indicou a maior produtividade para a razão mássica 1:1 entre as fontes de carbono (glicose) e nitrogênio (extrato de leveduras). O suprimento de oxigênio ao meio de cultura foi o principal fator para a produção do AH com as propriedades desejadas. A melhor produção foi 3,75 g/L, obtida a 2,0 vvm e agitação 150 rpm em 2,5 L de meio. O AH produzido apresentou viscosidade 913 cP, módulo elástico 73,1 Pa e massa molar 3,2 x 106 Da. O corte na alimentação de oxigênio causou despolimerização reduzindo a viscosidade e viscoelasticidade do AH. A determinação do coeficiente volumétrico de transferência de oxigênio permitiu caracterizar o suprimento adequado de oxigênio ao longo da fermentação. Os resultados obtidos demonstram a potencialidade da produção do AH por fermentação submersa e contribuem para o entendimento dos mecanismos envolvidos na sua formação e polimerização / Abstract: This work studied the production of hyaluronic acid (HA) by microbial route correlating variables of the process with productivity and physico-chemical properties of the product. Strains of Streptococcus were used in batch fermentations, aiming to obtain HA with high molecular weight, in the range o millions, high viscosity and viscoelasticity. Those properties are requested for medical applications of HA in the treatment of degenerative and inflammatory diseases of joints, as well as in the replacement of the sinovial fluid and as vehicle for drug delivery in surgical implants. Initially, the performance of three strains of mutants of Streptococcus (S. equi, ATCC 39506 and 6580, and S. zooepidemicus ATCC 39920), grown in medium of culture composed of glucose, yeast extract and salts were evaluated. The largest production of HA with viscous properties and viscoelasticity was obtained with the strain ATCC 39920. Secondly, a complete factorial planning of experiments 22 using the selected strain was performed aiming optimization the culture medium. The largest productivity was obtained for 1:1 (w/w) ratio between the sources of carbon (glucose) and nitrogen (yeast extract). The supplying of oxygen to the culture medium was the limiting factor to the production of AH with the required properties. The best production obtained was 3.75 g/L, using 2.0 vvm and 150 rpm agitation in 2.5 L medium. The AH produced had viscosity 913 cP, elastic module 73.1 Pa and molecular weight 3.2 x 106 Da. The interruption in the feeding of oxygen reduced the viscosity and viscoelasticity of HA, due to depolymerization. The determination of the volumetric coefficient for oxygen transfer allowed to detect changes in medium along fermentation. The obtained results demonstrate the potentiality of the production of HA by submerged fermentation and also contribute to the understanding of the mechanisms involved in HA formation and polymerization / Doutorado / Desenvolvimento de Processos Biotecnologicos / Doutor em Engenharia Química
97

Expressao, purificacao e caracterizacao de tireotrofina recombinante humana (rec-hTSH) em celulas de ovario de hamster chines (CHO)

MENDONCA, FERNANDA de 09 October 2014 (has links)
Made available in DSpace on 2014-10-09T12:48:54Z (GMT). No. of bitstreams: 0 / Made available in DSpace on 2014-10-09T14:01:11Z (GMT). No. of bitstreams: 1 09617.pdf: 5595560 bytes, checksum: 5a054c653d88ec87b2d3c979b79b00b5 (MD5) / Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP) / Dissertacao [Mestrado] / IPEN/D / Instituto de Pesquisas Energeticas e Nucleares - IPEN/CNEN-SP / FAPESP:00/09008-4
98

Estudo da polimerizacao do acetato de vinila utilizando a radiacao ionizante

MESQUITA, ANDREA C. 09 October 2014 (has links)
Made available in DSpace on 2014-10-09T12:47:50Z (GMT). No. of bitstreams: 0 / Made available in DSpace on 2014-10-09T14:08:26Z (GMT). No. of bitstreams: 1 08511.pdf: 4385457 bytes, checksum: a0c4d3d31c582b2a850f8616fec84e9e (MD5) / Dissertacao (Mestrado) / IPEN/D / Instituto de Pesquisas Energeticas e Nucleares - IPEN/CNEN-SP
99

Static and Flow Properties of Dilute Polymer Solutions

Whang, Kyu-ho 08 1900 (has links)
Small weight percentages of certain high-molecular weight polymers added to liquids in turbulent flow through conduits can result in dramatic friction reduction. Although many current and potential uses of the drag reduction phenomenon exist, there is a fundamental problem: drag reduction efficacy decreases rapidly with flow time due to the mechanical degradation in flow of the added polymer. In this thesis study, dilute aqueous solutions of polyacrylamide were tested under turbulent flow conditions in an attempt to determine where mechanical degradation in flow occurs.
100

An investigation of the effects of high molecular weight glutenin subunits on wheat tortilla quality

Pierucci, Valquiria Resende Malaspina January 1900 (has links)
Master of Science / Food Science Institute, Agriculture / Katherine A. Tilley / Michael Tilley / The wheat tortilla is a chemically leavened circular light colored flat bread. Desirable characteristics for good quality tortilla include large diameter, softness, flexibility and long shelf stability. Important components influencing quality are wheat flour properties, which have not been optimized for tortilla industrial production thus far. The studies presented here investigated the effects of high molecular weight glutenin subunits (HMW-GS) on tortilla quality. Two approaches were employed: biotypes derived from Centurk and OK102 cultivars expressing defined HMW-GS compositions and transgenic wheat lines over-expressing HMW-GS 10. Analysis of protein expression and protein extractability were conducted to characterize wheat flours and suitable assays carried out to determine the respective dough properties. Tortillas were prepared by the hot-press method and quality parameters were measured at days 0, 2, 4, 7 and 14. Tortillas derived from Centurk biotypes possessing HMW-GS 2*, 7+9, 2+12, 2*, 7+8, 5+10 and 2*, 7+9, 5+10 exhibited superior texture profiles over time, but smaller diameters than the biotype 2*, 7+8, 2+12. Tortillas containing HMW-GS 7+9 and 2+12 revealed a texture profile similar to tortillas containing 5+10. Tortillas from the OK biotype 2*, 7+9, 3+12 exhibited larger diameter and texture profiles equivalent to tortillas containing 5+10. Therefore, this biotype showed the best quality within this cultivar. Tortillas derived from transgenic flours over-expressing HMW-GS 10 exhibited an undesirable rough appearance with decreased diameter, greater thickness, lower rollability scores, lower stretchability and greater rupture force over time. Over-expression of HMW-GS 10 in a wheat line containing 1RS-translocation did not promote the same deleterious effects in tortilla quality as it did in transgenic lines without 1RS translocation.

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