5,274 research outputs found
Erratum: Update on psychological functioning in adults with congenital heart disease: A systematic review (Expert Review of Cardiovascular Therapy (2013) 11(6) (785–791) (10.1586/erc.13.9))
No full textThe correct author affiliations are given below: Edward Callus*1, Emilia Quadri1, Cristian Ricci2, Cristiana Passerini1, Anna Tovo1, Gabriele Pelissero3 and Massimo Chessa11IRCCS Policlinico San Donato, Pediatric and Adult Congenital Heart Centre, Italy 2Fondazione Don C. Gnocchi, Italy 3IRCCS Policlinico San Donato, Scientific Directorate, Italy *Author for correspondence: Tel.: +39 02 527 747 07 [email protected]. © 2013, Informa UK, Ltd. All rights reserved
Cultura de calos e criopreservação de sementes de Cedrela fissilis Vellozo (Meliaceae)
No full textDissertação (mestrado) - Universidade Federal de Santa Catarina. Programa de Pós-Graduação em BiotecnologiaCedrela fissilis Vellozo (Meliaceae) é uma espécie nativa da Floresta Atlântica, que produz madeira de grande valor econômico. Os objetivos deste trabalho foram estudar o crescimento de calos em diferentes condições de cultivo, realizar análises preliminares do teor de óleos essenciais e dos constituintes voláteis produzidos pelos mesmos e desenvolver estudos sobre a criopreservação de sementes e conservação in vitro de propágulos vegetativos. Em todos os experimentos foram utilizados segmentos nodais cotiledonares, exceto para testar o efeito de diferentes tipos de explantes, em que segmentos apicais e foliares também foram utilizados. O meio de cultura Murashige & Skoog foi utilizado em todos os experimentos, suplementado com 2,5 M de 6-benzilaminopurina (BAP) e 5 M de ácido a-naftalenoacético (ANA), 0,2% de Phytagel e com 58,4 mM de sacarose, exceto nos casos em que outras fontes de carbono foram testadas ou em que foram testadas diferentes concentrações de sacarose na ausência ou na presença de glutamina ou em que diferentes concentrações de BAP e ANA foram utilizadas separadamente ou em combinação. As culturas foram mantidas em sala de crescimento com temperatura controlada de 25 2ºC, sob fotoperíodo de 16 horas (exceto as submetidas ao escuro), provido por lâmpadas fluorescentes Philips TDL (22.3 µmol.m-².s-¹) e foram avaliadas, após quatro e oito semanas, dependendo do experimento, através da determinação da massa fresca, massa seca e do teor de água. Os óleos essenciais foram extraídos por hidrodestilação de calos crescidos na luz em 2,5 M de BAP e 5 M de ANA . A extração dos constituintes voláteis foi realizada pelo método líquido-sólido e análise através de cromatografia gasosa. As sementes criopreservadas foram descongeladas em temperatura ambiente ou a 45 C, em banho-maria, por três minutos. Os segmentos nodais foliares foram armazenados, por seis semanas em meio de cultura Murashige & Skoog, com metade da concentração salina, desprovido de sacarose, a 25 C e a 5 C e após o período de armazenamento transferidos para o meio de cultura Murashige & Skoog, suplementado com 2,5 M de BAP e 5 M de ANA. Nenhuma diferença foi detectada entre os valores de massa fresca dos calos iniciados a partir de segmentos nodais cotiledonares e apicais e os maiores valores foram obtidos com 2,5 M de BAP e 5 M de ANA. Não foram detectadas diferenças entre os maiores valores em massa seca de calos iniciados a partir de segmentos nodais cotiledonares e foliares, obtidos com oito semanas de cultivo. Os calos iniciados a partir de segmentos nodais cotiledonares cresceram em todas as combinações de ANA e BAP testadas. Os maiores valores de massa seca foram obtidos em 2,5 M de ANA com 2,5 M de BAP, 5 M de ANA e 2,5 M ou 5 M de BAP. A maior massa seca dos calos ocorreu após oito semanas de cultivo em sacarose, sendo os valores observados para glicose e frutose semelhantes. Os maiores valores de massa seca dos calos foram obtidos com 58,4 mM ou 116,8 mM de sacarose e com 87,6 mM de sacarose e 2,73 mM de glutamina. Os calos apresentaram maior massa seca na luz, com oito semanas de cultivo e maior teor de água no escuro. Calos produzidos a partir de plântulas de diferentes idades não diferiram em massa seca, apenas no teor de água, que foi maior para os calos obtidos de explantes de plântulas com cinco semanas de idade. O rendimento de óleos essenciais extraídos por hidrodestilação, produzidos pelos calos foi de 1,17 % da massa seca. O linalol foi o composto majoritário produzido por calos crescidos na luz ou no escuro. Sete compostos voláteis foram identificados em calos crescidos no escuro e doze foram identificados em calos crescidos na luz. O germacreno apareceu em maior proporção nos calos crescidos na luz. A massa seca dos calos produzidos a partir de segmentos nodais foliares armazenados por seis semanas a 5 C foi maior do que a massa seca dos calos produzidos pelos segmentos apicais, não havendo diferença entre as massas secas quando ambos os explantes foram armazenados a 25 C. As sementes toleraram a criopreservação por quinze meses e o descongelamento lento. O estabelecimento de diferentes condições de cultivo de calos, a detecção de compostos de interesse nos mesmos e o estabelecimento de métodos que permitem a criopreservação, por longos períodos de tempo, e a conservação in vitro, por curtos períodos de tempo, de propágulos vegetativos indicam o potencial da aplicação de abordagens biotecnológicas para a produção de metabólitos secundários e conservação de germoplasma desta espécie. Cedrela fissilis Vellozo (Meliaceae) is a native species of the Atlantic Forest, important for timber production. The aim of this work is to study callus growth in different culture conditions, to carry out preliminary analysis on the content of essential oils and its volatile components and to develop studies on seed cryopreservation and in vitro conservation of vegetative propagules. Cotyledonary nodal segments were used in all experiments, except when different type of explants were used. The Murashige & Skoog medium supplemented with 2,5 M 6-benzilaminopurine (BAP) and 5 M a-naphtaleneacetic acid (ANA), 0,2% Phytagel and 58,4 mM sucrose was used in all experiments, except when different carbon sources, different sucrose concentration, in the absence or presence of glutamine or different concentration of BAP and ANA, isolated or combined, were tested. The cultures were maintained under 25 2ºC, 16 hours photoperiod (except those kept in the dark), provided by fluorescent Philips TDL (22.3 µmol.m-².s-¹) tubes and were assessed after either four or eight weeks, depending on the experiment, for fresh mass, dry mass and water content. The essential oils were extracted by hydrodistillation from callus grown under light in 2,5 M BAP and 5 M ANA. The extraction of volatile components was carried out through the liquid-solid method and the analysis by gas chromatography. Seeds cryopreserved in liquid nitrogen and thawed either at room temperature or at 45 C, in a water bath, for three minutes. Leaf nodal segments and apical shoot segments were stored at either 25 C or 5 C, for six weeks in half strength Murashige & Skoog medium, deprived of sucrose, and transferred, after storage, to the Murashige & Skoog medium supplemented with 2,5 M BAP and 5 M ANA. No differences were detected in the fresh mass of calluses initiated either from cotyledonary nodal segments or apical segments and the highest values were obtained in the culture medium with 2,5 M BAP and 5 M ANA. No differences were detected in the highest dry mass values of calluses initiated from either cotyledonary nodal segments or leaf nodal segments, after eight weeks of culture. Growth of calluses initiated from cotyledonary nodal segments was achieved in all combination of ANA and BAP tested. Superior values of dry mass were obtained in 2,5 M ANA with 2,5 M BAP, 5 M ANA and 2,5 M or 5 M BAP. The highest callus dry mass occurred after eight weeks of culture in sucrose, and similar values were obtained for glucose and fructose. Superior values of callus dry mass were obtained on either 58,4 mM or 116,8 mM sucrose and on 87,6 mM sucrose and 2,73 mM glutamine. Calluses showed higher dry mass in the light, when cultivated for eight weeks. Calluses produced from seedlings with different age did not differ in dry mass, only the water content was higher in callus obtained from explants of five week-old seedlings. The content of essential oils produced by the callus was 1,17 % of the dry mass. Linalool was the major compound produced by callus grown either under light or in the dark. Seven volatile compounds were identified in callus grown in the dark and twelve were identified in callus grown in the light. Germacrene appeared in higher proportion in the callus grown in the light. The dry mass of calluses produced from leaf nodal segments stored for six weeks at 5 C was higher than the dry mass of calluses produced from the apical segments, and no difference in the dry mass was detected when both explants were stored at 25 C. Seeds tolerated cryopreservation for fifteen months and the slow thawing. The establishment of different callus culture condition, the detection of important compounds produced by the callus and the establishment of methods which allow long term seed cryopreservation, and short term in vitro conservation of vegetative propagules indicate the potential of application of biotechnological approaches for the production of secondary metabolites and germplasm conservation of this species
Influence of the mechanical environment upon the healing of segmental bone defects in the rat femur
No full textLoss of large segments of bone creates critical size defects (CSDs). These fail to heal
spontaneously and present major clinical challenges to orthopaedic surgeons. The
research described in this thesis is based upon the hypothesis that the healing of
CSDs is responsive to the ambient mechanical environment, and can be accelerated
by mechanical modulation. This hypothesis was tested in rat, femoral CSDs treated
with recombinant, human, bone morphogenetic protein-2.
For this study I designed novel external fixators allowing experimental control over
the local mechanical environment. These were characterised by extensive
mechanical testing prior to evaluation in the rat model.
Low stiffness fixators induced callus formation 9 days after surgery, whereas rigid
fixation delayed it until 2 weeks. All defects were radiologically bridged after 3
weeks. Rats were euthanised after 8 weeks and the defects evaluated by a battery of
imaging, mechanical and histological tests. All confirmed the superiority of the
lowest stiffness fixators.
Based upon these data, I hypothesised that healing would be improved by imposing
low stiffness for the first two weeks of healing, followed by high stiffness for the
remaining six weeks. The experimental data confirm that this regimen dramatically
accelerated callus formation and maturation, and induced faster remodelling of
endosteal and periosteal callus. This was associated with higher failure strength,
fewer trabeculae, decreased callus size and thicker and more uniform distribution of
new cortical bone. Histologically it was not possible to detect cartilage within the
defects prior to the appearance of bone, suggesting that healing either does not occur
through endochondral ossification, or that this process is very rapid.
These data confirm that the healing of CSDs is highly responsive to the ambient
mechanical environment, allowing the rate and quality of healing to be manipulated.
This information will help develop more efficient ways to heal CSD clinically
Calos de passifloras silvestres: indução, caracterização morfo-histológica, metabólica e ultraestrutural
No full textDissertação (mestrado) - Universidade Federal de Santa Catarina, Centro de Ciências Biológicas, Programa de Pós-Graduação em Biologia de Fungos, Algas e Plantas, Florianópolis, 2015.Passiflora setacea e P. tenuifila são espécies silvestres de maracujás endêmicas do Brasil, com importantes propriedades medicinais. A P. setacea apresenta também potencial para o mercado consumidor devido à qualidade de seus frutos e a P. tenuifila tem sido indicada para o melhoramento genético. A produção de metabólitos secundários via cultura de tecidos in vitro tem sido investigada como uma alternativa à utilização de plantas produzidas no campo, pois oferece ferramentas para promover a otimização da produção. Assim, os objetivos deste estudo foram: analisar a histologia de calos de P. setacea induzidos em 2,4-D; determinar efeitos de diferentes concentrações de ANA e fontes de Carbono, quanto à morfo-histologia, ultraestrutura e teores de metabólitos de calos de P.tenuifila e; caracterizar, com base na ultraestrutura, como a Matriz Extracelular (ME) determina a desintegração das células dos calos friáveis de espécies de Passilfora. Para o estudo sobre indução e histologia, amostras de calos de P. setacea e P. tenuifila, obtidos a partir de diferentes tipos de explantes e meios de cultivo in vitro, foram fixadas em glutaraldeído 2,5% com tampão fosfato de sódio 0,1M, pH 7,2 e desidratadas. Para MO, amostras foram infiltradas em hidroxietilmetacrilato, seccionadas em micrótomo, coradas com azul de toluidina ou submetidas à reação com vermelho de rutênio, para identificar substâncias pécticas. Para análise em MEV, amostras foram secas em ponto crítico de CO2 e, para análise em MET, amostras de calos de P.tenuifila foram pós-fixadas em tetróxido de ósmio, infiltradas em resina Spurr, seccionadas em ultramicrótomo, contrastadas com Acetato de Uranila e Citrato de Chumbo. Calos de P. tenuifila com 45 dias foram coletados para determinar: percentagem de explantes que induziram calos, teor de água, acúmulo de biomassa, análise de metabólitos, atividade antioxidante por DPPH e a dosagem de clorofila. A percentagem mais elevada de explantes que formaram calos de P. setacea foi observada em 2,5 e 5,0 µM de 2,4-D. Calos obtidos a partir de segmentos de hipocótilo em 5,0 µM de 2,4-D e de segmento nodal foliar em 2,5 µM de 2,4-D mostraram características histológicas indicando grande potencial para morfogênese e produção de metabólitos secundários. Em estudo com calos de P. tenufila, todas as fontes de carbono e concentrações de ANA permitiram altas22percentagens de produção de calos, sem diferenças entre os tratamentos. Todos os calos apresentaram textura friável e características histológicas similares. Compostos fenólicos, flavonóides e carotenóides (luteína, zeaxantina e a-caroteno) foram detectados em calos de P. tenuifila. Atividade antioxidante não foi registrada. As características histológicas observadas sugerem que os calos de P.tenuifila, com 45 dias de cultivo, estavam em estádio inicial. No estudo sobre a ME verificamos sua constituição péctica. A superfície dos calos estava revestida com ME membranosa, com aspecto liso externamente e granular ou fibrilar internamente. O aspecto granular ou fibrilar decorre da desconexão entre células. A presença de golgi, vesículas na periferia da célula e evidências da liberação de substâncias e dissociação intercelular foram registradas ultraestruturalmente e sugerem que a consistência friável dos calos de Passiflora é resultante da desconexão das células periféricas.Abstract : Passiflora setacea and P. tenuifila are wild species of endemic passion fruit in Brazil, with important medicinal properties. P. setacea has also potential for the consumer market due to the quality of its fruit and P. tenuifila has been used for the genetic improvement. The production of secondary metabolites through in vitro tissue culture has been investigated as an alternative to the use of plants produced in the field, it provides tools for promoting the production optimization. The objectives of this study were to analyze the histology of P. setacea callus induced in 2,4-D; examine the effects of different concentrations of NAA and carbon sources in the morphological- histology, ultrastructure and level of metabolites in P.tenuifila callus and; to characterize, based on ultrastructure, how the Extracelular Matrix (EM) determines the disintegration of cells in friable callus of Passilfora species. For the study of induction and histology, samples of P. tenuifila and P. setacea callus, obtained from differents types of explants and in vitro culture media were fixed in 2.5% glutaraldehyde in sodium phosphate buffer 0, 1M, pH 7.2 and dried. For OM, the samples were infiltrated in methacrylate, sectioned with a microtome, stained with toluidine blue or subjected to reaction with ruthenium red to identify pectic substances. For SEM analysis, samples were dried in a critical point of CO2, and for TEM analysis, samples P.tenuifila callus were post-fixed in osmium tetroxide, infiltrated in Spurr resin, sectioned with ultramicrotome, contrasted with uranyl acetate and lead citrate. P.tenuifila calllus were collected with 45 days to determine: percentage of explants that to induced callus, water content, biomass accumulation, analysis of metabolites, antioxidant activity and measurement of chlorophyll. The highest percentage of explants that to induced P. setacea callus was observed at 2.5 and 5.0 uM of 2,4-D. Callus from hypocotyl segment in 5.0 µM 2,4-D and from leaf nodal segment in 2.5 µM 2,4-D showed histological characteristics indicate great potential for morphogenesis and secondary metabolite production. In the study with P. tenufila callus, all sources of carbon and NAA concentrations allowed high induction percentages, without differences between treatments. All callus showed friable texture and similar histological characteristics. Phenolic compounds, flavonoids and carotenoids (lutein, zeaxanthin and a-carotene) were detected in P. tenuifila callus.24Antioxidant activity wasn`t recorded. The histological characteristics suggest that P.tenuifila callus with 45 days were in inicial stage. In the study about the EM we found your pectic constitution. The surface of the callus was coated with EM membranous, with smooth appearance externally and granular or fibrillar internally. The granular or fibrillar aspect stems from the disconnect between cells. The presence of Golgi, vesicles at the periphery of the cell and evidence of release of substances and intercellular dissociation were recorded by ultrastructure study and suggest that the Passiflora friable callus is the result of disconnection of peripheral cells
Perfis metabílocos de calos de Cedrela fissilis Vell. (Meliaceae) cultivados in vitro
No full textDissertação (mestrado) - Universidade Federal de Santa Catarina, Centro de Ciências Biológicas, Programa de Pós-graduação em Biologia Vegetal, Florianópolis, 2013Abstract: Cedrela fissilis Velloso (Meliaceae) is a fast growing native tree from the Atlantic Forest economically important for its medicinal properties and for producing valuable hardwood. The objectives of this work were to study the effects of carbon sources (sucrose, glucose, fructose), glutamine, explant types (leaf node, leaf, epicotyl, cotyledonary node, cotyledon, hypocotyl, root), growth regulators (BAP and NAA), light and dark on the levels of total proteins, total soluble sugars, starch, total phenolics, flavonoids, carotenoids (content and types), chlorophylls a and b, metabolic profiles, antioxidant activity of in vitro cultured calluses, and to evaluate the morphology and hystochemistry of callus originated from the different explants type cultured on glucose. Calluses were induced on Murashige and Skoog culture medium supplemented with 0,2% Phytagel from all types of explant tested (except from leaf explant cultured on 118 mM sucrose, 2,73 mM glutamine in the light), cultured on different carbon sources (sucrose, fructose, glucose), in all combinations of BAP and NAA concentration (2,5 µM e 5 µM), in different glutamine concentration (0; 2,73 e 5,46 µM), either in light or in the dark. The manipulation of sucrose (59 mM e 118 mM) and glutamine concentration (0; 2,73 e 5,46 mM) in Murashige & Skoog, supplemented with 0,2% de Phytagel, 2,5 µM BAP and 5 µM NAA allowed the establishment of optimal callus culture condition for cotyledonary node which promoted the production of the highest levels of total proteins (59 mM sucrose and 5,46 mM glutamine), total soluble sugars (118 mM sucrose and 5,46 mM glutamine), starch (118 mM sucrose and 5,46 mM glutamine), total phenolics (118 mM sucrose and 2,73 mM de glutamine), flavonoids (118 mM sucrose and 2,73 mM glutamine), carotenoids (118 mM sucrose and 5,46 mM glutamine), chlorophylls a (59 mM sucrose and 5,46 mM de glutamine) e b (59 mM de sucrose and 5,46 mM glutamine, 118 mM sucrose and 2,73 mM glutamine) and highest percentage of DPPH free radical inhibition (118 mM sucrose and 2,73 mM de glutamine).The manipulation of carbon sources (118 mM sucrose, fructose, glucose) in combination with BAP and NAA (2,5 µM e 5 µM), in MS culture medium supplemented with 0,2% de Phytagel and 2,73 mM glutamine optimized the biosynthesis of total proteins (sucrose, 5 µM BAP and 5 µM NAA; fructose 2,5 µM de BAP and 5 µM NAA), total soluble sugars (sucrose, 2,5 µM BAP and 2,5 µM NAA), starch (sucrose, 2,5 µM BAP and 2,5 µM NAA), total phenolics (sucrose, 2,5 µM BAP and 2,5 µM NAA) , flavonoids (glucose 2,5 µM BAP and 2,5 µM NAA), carotenoids (sucrose, 5 µM BAP and 2,5 µM NAA), chlorophylls a (glucose 5 µM BAP and 2,5 µM NAA) and b (sucrose, 2,5 µM BAP and 2,5 µM NAA; glucose 5 µM BAP and 2,5 µM NAA) and the highest percentage of DPPH free radical inhibition (glucose 5 µM BAP and 5 µM NAA) in callus originated from cotyledonary nodes.The manipulation of explant types (leaf node, cotyledonar node, segments of epicotyl, leaf, cotyledon, hypocotyl, root), carbon source (118 mM sucrose, fructose, glucose), glutamine (0; 2,73 mM), light and dark promoted the biosynthesis of total proteins (sucrose, cotyledon, without glutamine, light), total soluble sugars (sucrose, cotyledon, leaf node, without glutamine, light), starch (sucrose, leaf node, without glutamine, light), total phenolics (sucrose, cotyledon, glutamine, light), flavonoids (sucrose, cotyledon, glutamine, light; fructose, leaf, without glutamine, light), carotenoids (fructose or glucose, leaf, glutamine, light), chlorophyll a (sucrose, cotyledon, glutamine, light; fructose, leaf, cotyledon, without glutamine, light; glucose, leaf, glutamine light); chlorophyll b (sucrose, cotyledon, glutamine, light; fructose, leaf, without glutamine, light; glucose, leaf, glutamine light); the percentage of DPPH free radical inhibition higher than 70% (sucrose, all explants types cultured either with or without glutamine, in light or dark, except for cotyledonary node (with glutamine, light), hypocotyls (with glutamine, light or without glutamine, dark).The carbon sources, glutamine, BAP and NAA influenced the calluses metabolic profiles determined through the UV-vis sacanning. The carbon sources, explants types and glutamine affected the types and concentrations of carotenoids produced by calluses, fructose promoted the biosynthesis of carotenoids types as beta-cryptoxantin and trans-betacryptoxantin which were not produced with sucrose or were detected in a few treatments with glucose, as demonstrated by the HPLC analysis. The AT-O, PAS e CBB tests indicated, respectivelly, the metacromatic reaction in the callus cell wall, the presence of ortocromatic granules, possibly phenolics and positive reactions for neutral polyssacarides, as cellulose. Few starch grains and rich protein organels were detected in the cytoplasm. The results obtained in this study showed that optimal callus culture conditions were established, through the manipulation of carbon sources, explant type, glutamine, light and dark, which increased the production of a range of metabolites of interest and could foster further investigation on biosynthesis, identification and optimization of the production of important compounds by the cell culture systems developed
Crescimento, perfil metabólico e citoquímica de calos de Cedrela fissilis Vellozo (Meliaceae)
No full textDissertação (mestrado) - Universidade Federal de Santa Catarina, Centro de Ciências Biológicas, Programa de Pós-Graduação em Biotecnologia, Florianópolis, 2011Cedrela fissilis Velloso (Meliaceae) é uma árvore nativa da Floresta Atlântica (Sul do Brasil) de grande interesse econômico devido à sua madeira de qualidade e seu crescimento rápido, ideal para programas de reflorestamento e arborização de espaços urbanos. Assim como as demais espécies da Família Meliaceae, seu metabolismo secundário é caracterizado pela produção de terpenos, que tem despertado grande interesse industrial e medicinal pelas suas atividades inseticida, anti-tumoral, antibiótica, anti-viral, anti-fúngica e anti-malárica. No entanto, as áreas de Floresta Atlântica têm sido suprimidas pela expansão das cidades e da agropecuária no Brasil, colocando em risco toda a biodiversidade deste ecossistema e tornando inviável a exploração sustentável de seus recursos. A cultura vegetal in vitro, através de técnicas de cultura de calos e suspensões celulares, é uma alternativa viável que permite a produção de compostos vegetais de interesse econômico sem que haja a necessidade da exploração dos recursos vegetais in loco, permitindo a conservação das florestas. Assim, este trabalho teve como objetivos estudar o efeito de diferentes condições de cultivo sobre o crescimento, o perfil metabólico e a histologia de calos de C. fissilis. O regulador de crescimento 2,4-D promoveu o crescimento de calos a partir de segmentos nodais cotiledonares e de hipocótilo com uma frequência entre 53,8 # 97,1%, porém demonstrou ser tóxico para segmentos foliares e cotiledonares. Os meios de cultura contendo glucose ou frutose promoveram melhor indução de calos do que os meios de cultura suplementado com sacarose. Calos crescidos na presença de glucose ou frutose formaram maior massa fresca e possuíam elevados teores de água. No entanto, calos crescidos em meio de cultura contendo sacarose formaram maior quantidade de massa seca. Diferentes combinações de BAP e ANA foram testadas e todas apresentaram elevados índices de crescimento de calos na presença de glucose ou frutose no meio de cultura. No meio de cultura contendo sacarose, a calogênese variou entre 48,3 # 86,7%, sendo que a combinação de 5 µM de BAP com 2,5 µM de ANA apresentou a maior taxa de calogênese. Foi observado que a posição do explante interfere na calogênese, e os segmentos nodais cotiledonares inoculados na posição vertical apresentaram maior frequência de calogênese (86,7%) do que os explantes inoculados na posição horizontal (63,3%). Segmentos nodais cotiledonares, segmentos apicais e segmentos de hipocótilo formaram calos em maior frequência e com maior massa fresca e seca. Segmentos de raízes formaram calos com menor massa seca, e segmentos foliares e cotiledonares formaram elevadas taxas de calos com raízes e com menor massa seca. O cultivo dos calos no escuro aumentou a frequência de calogênese em todos os tipos de explantes, mas provocou uma redução de massa seca. O aumento da concentração de glucose, frutose e sacarose no meio de cultura e do período de cultivo dos calos resultaram em maiores quantidades de massa fresca e seca. A adição de glutamina ao meio de cultura teve efeito inibitório sobre a indução e o crescimento dos calos. Sete formulações salinas de meio de cultura foram testadas, mas apenas no meio de cultura MS foi observava a calogênese. As curvas de dissimilação demonstraram que o crescimento dos calos na presença de frutose ocorre de maneira mais rápida e atinge o estágio estacionário antes dos calos crescidos em meio de cultura contendo glucose ou sacarose, e que a adição de glutamina ao meio de cultura, indiferentemente do açúcar utilizado, aumenta a fase lag e reduz o crescimento dos calos. A análise de varredura em UV-vis apontou a presença de compostos fenólicos, clorofilas e carotenóides nos calos e permitiu comparar a ocorrência destes metabólitos entre diferentes tratamentos. A dosagem de metabólitos secundários demonstrou que os calos crescidos em meio de cultura contendo frutose possuíam o maior teor de clorofila (116 µg/g MF), e que os calos crescidos em meio de cultura contendo sacarose e glutamina possuíam o maior conteúdo de compostos fenólicos totais (17,9 µg/g MS) e de flavonóides (2,89 µg/g). A espectroscopia por FTIR permitiu comparar o perfil metabólico primário e secundário de calos crescidos em diferentes condições de cultivo, evidenciando diferenças quali-quantitativas a nível molecular que podem ajudar a caracterizar os diferentes tratamentos avaliados. Na análise histoquímica de diferentes morfologias de calos foi possível observar que as células possuem grandes vacúolos e que ocorre a formação de arranjos concêntricos de células que caracterizam o início da formação de órgãos. A coloração com PAS reagiu com os carboidratos neutros da parede celular e com os grânulos de amido no citoplasma das células, presentes em grande quantidade nos calos com morfologia nodular. A coloração de AT-O teve apenas uma leve reação metacromática na parede celular das células, indicando pouca ocorrência de açúcares ácidos. A coloração de CBB tingiu de maneira uniforme o citoplasma das células devido à presença de proteínas livres e em organelas e destacou a presença de alguns grânulos protéicos. A calogênese é influenciada pela composição do meio de cultura, condições ambientais de cultivo e por fatores endógenos, como o balanço hormonal de cada tecido e o genótipo da planta doadora dos explantes. As curvas de dissimilação, a análise de metabólitos primários e secundários através de espectroscopia de UV-vis e FTIR e o conhecimento da organização celular através da histoquímica fornecem suporte para a otimização das culturas de calos e para a identificação e seleção de culturas com potencial para a produção de compostos de interesse.Cedrela fissilis Velloso (Meliaceae) is a fast growing timber tree from Atlantic Forest (South Brazil) which has great economic value due to its high quality wood and its uses in reforestation programs and landscaping of urban areas. As other Meliaceae species, its secondary metabolism is characterized by the production of terpenes of industrial and medicinal interests for its insecticidal, anti-tumor, antibiotic, anti-viral, anti-fungal e anti-malarial activities. However, Atlantic Forest areas have been suppressed by urban and agriculture expansion, which endangers the biodiversity of this ecosystem and make unfeasible the sustainable exploitation of natural resources. In vitro plant tissue cultures, through the use of callus and cell suspension cultures, is a practicable alternative that allows the production of plant compounds of economic interest without the need of plant resources in loco, allowing forests conservation. This work aimed the study the effects of different culture conditions on growth, metabolic profile and histology of C. fissilis callus cultures. The plant growth regulator 2,4-D promoted callus growth from cotyledonary nodal segments and hypocotil segments with a frequency between 53,8 # 97,1%, but showed to be toxic to cotyledon and leaf segments. Culture media containing glucose or fructose promoted higher callus formation than culture medium containing sucrose. Callus growth in the presence of glucose or fructose formed superior fresh weight and had higher water content. In the other hand, callus growth on culture media containing sucrose formed higher dry matter. Different combinations of BAP and ANA were tested and all of them promoted callus growth in culture media containing glucose or fructose. In the presence of sucrose, callus formation ranged between 48,3 # 86,7%, and the combination of BAP 5 ìM with ANA 2,5 ìM induced the best calogenesis frequency. The explant position interfered on callus formation, and the cotyledonary nodal segments inoculated in the vertical position promoted higher callus induction (86,7%) than those inoculated in the horizontal position (63,3%) but no differences on callus growth were observed. Cotyledonary nodal segments, apical segments and hypocotil segments formed callus in a higher frequency and with major fresh and dry matter. Callus from root segments formed less dry matter, and cotyledon and leaf segments formed callus and roots. The increase of sugar concentration in the culture medium and of the culture period promoted the increase in fresh and dry matter. The addition of glutamine to the culture media inhibited callus induction and growth. Seven culture media formulas were tested, but callus formation occurred only in the MS culture medium. Dissimilation curves showed that callus growth is faster in culture medium containing fructose, but cultures reach stationary period earlier. The addition of glutamine to the culture media increased the lag phase and reduced callus growth. The UV-vis screen showed the presence of phenolic compounds, chlorophyll and carotenoids in callus cells and allowed the comparison of these metabolites between calluses from different treatments. The secondary metabolites quantification showed that callus grown in the presence of fructose had the higher chlorophyll content (116 ìg/g FM), whilst callus growth in the presence of sucrose and glutamine had higher phenolic compounds (17,9 ìg/g DM) and flavonoids content (2,89 ìg/g). FTIR spectroscopy allowed the comparison between primary and secondary metabolic profile of calluses grown under different culture conditions and showed qualitative and quantitative differences. The histochemical analysis of different callus morphologies showed cells containing large vacuoles and concentrically cells arrangements that characterizes first stages of organ formation. PAS staining has reacted with neutral carbohydrates of cell walls and starch grains in cells cytoplasm, which were numerous in nodular callus morphology. TB-O staining had a weak metachromatic reaction with cell walls, indicating that acid carbohydrates occurs in low quantity in callus cells. CBB staining reacted with cytoplasm proteins and organelles and with few protein grains. Callus inductions and growth are determined by culture medium composition, environmental conditions and endogenous factors, such as hormonal balance and genotype. Dissimilation curves, primary and secondary metabolites analysis through UV-vis and FTIR spectroscopy, and the study of callus cell organization by histochemistry provided the background for further studies on callus culture optimization and for identification and selection of cultures with potential to produce compounds of economic interest
µCT callus analysis: (A) callus mineral content, (B) callus volume, (C) callus mineral density.
No full text<p>Pten mutants had more callus mineral content, volume and density at each time point. (*<i>p</i><0.05 WT to Mut at the time point.).</p
Sequencing statistics of Se-treated and untreated <i>A</i>. <i>chrysochlorus</i> callus tissues.
No full text<p>Sequencing statistics of Se-treated and untreated <i>A</i>. <i>chrysochlorus</i> callus tissues.</p
Increased regeneration efficiency of _Brassica napus_ L. cultivars Star, Westar and Cyclone from hypocotyle and cotyledonary explants
No full textThe comparative organogenesis of _Brassica napus_ L cultivars Cyclone, Star and Westar was studied. The cotyledonary explants gave a higher response to all the combinations of 0.5 mg/L 2,4-D and BAP (0.5, 1.0,1.5 and 2.0 mg/L} used for optimizing the conditions for callus induction. The best mean weight and mean length of callus was obtained at 0.5 mg/L 2,4-D and 1.5mg/L BAP for Star cotyledonary explants. For the complete plant regeneration the new method of exposing the explants culture to Growth regulator free medium was performed. The method was applicable to both hypocotyl and cotyledonary explants. The Shoot Induction Frequency for hypocotyl (6-34%) in the three cultivars is higher than the cotyledonary explants (3-23%). The method is speedy and almost all of the shoots and some unshooted calli (78%) form roots on the same media without prior transfer to rooting medium
Microspore developmental stage and anther length influence the induction of tomato anther callus
No full textAnthers of L680A, Licato and Ailsa Craig tomato (Lycopersicon esculentum) were plated on Doy's basal medium 1 to determine whether microspore developmental stage and anther length influence anther callus production. Although calli were induced at all stages of anther development, anthers containing prophase I-stage microspores produced the highest frequency of calli. Fewer calli were produced as microspores approached the uninucleate and binucleate pollen stage. Callus diameter also decreased as anther development progressed. Significantly larger calli were produced from prophase I than later-stage anthers. Time of anther harvest (morning vs. afternoon) did not significantly affect callus number or diameter. Anther and flower bud length were both significantly correlated with anther developmental stage, the number of anthers producing calli, and mean calli diameter. In each case, anther length exhibited a significantly better correlation than bud length..RE: 20 ref.; SC: CA; HO; PL; 0C; 0P; 7BSource type: Electronic(1) http://upei-resolver.asin-risa.ca?sid=SP:CABI&id=pmid:&id=&issn=0018-5345&isbn=&volume=27&issue=7&spage=838&pages=838-840&date=1992&title=HortScience%20&atitle=Microspore%20developmental%20stage%20and%20anther%20length%20influence%20the%20induction%20of%20tomato%20anther%20callus.&aulast=Summers&pid=%3Cauthor%3ESummers%2c%20W%20L%3bJaramillo%2c%20J%3bBailey%2c%20T%3C%2Fauthor%3E%3CAN%3E19931642181%3C%2FAN%3E%3CDT%3EJournal%20article%3C%2FDT%3
- …
