Embriogênese somática em Araucaria angustifolia (Bert.) O. Kuntze: pontos de controle e aspectos do metabolismo /

Dissertação (mestrado) - Universidade Federal de Santa Catarina, Centro de Ciências Agrárias.Durante grande parte do século XX a Araucaria angustifolia (Bert.) O. Kuntze, (Araucariaceae), foi a espécie madeireira nativa mais importante do sul do Brasil. Em coníferas, a embriogênese somática destaca-se como uma alternativa viável para a conservação e melhoramento genético de espécies florestais. No presente trabalho, estudaram-se alguns pontos de controle do desenvolvimento embrionário em A. angustifolia, visando o estabelecimento de um protocolo para a obtenção de embriões somáticos. Culturas embriogênicas foram induzidas a partir de embriões zigóticos de sementes provenientes de três diferentes genótipos. Análises histoquímicas das culturas embriogênicas evidenciaram pró-embriões somáticos em diferentes estádios de desenvolvimento. Na fase de maturação, foram testados fontes e níveis de fitorreguladores: ABA, BAP e Kin ; agentes osmóticos PEG 3350, PEG 8000, BSA e inositol; maltose. Durante a fase de manutenção, foram realizados estudos sobre alguns parâmetros de crescimento de suspensões celulares de linhagens celulares embriogênicas. Analisou-se o crescimento das linhagens celulares, determinação dos valores de pH e concentração de glucose no meio de cultura e quantificação de proteínas extracelulares. Além disso, foi determinado o perfil protéico das linhagens celulares, e de embriões zigóticos e megagametófitos através de eletroforese em gel de poliacrilamida

Cloning and expression of embryogenesis-regulating genes in Araucaria angustifolia (Bert.) O. Kuntze (Brazilian Pine)

Angiosperm and gymnosperm plants evolved from a common ancestor about 300 million years ago. Apart from morphological and structural differences in embryogenesis and seed origin, a set of embryogenesis-regulating genes and the molecular mechanisms involved in embryo development seem to have been conserved alike in both taxa. Few studies have covered molecular aspects of embryogenesis in the Brazilian pine, the only economically important native conifer in Brazil. Thus eight embryogenesis-regulating genes, viz., ARGONAUTE 1, CUP-SHAPED COTYLEDON 1, WUSCHEL-related WOX, S-LOCUS LECTIN PROTEIN KINASE, SCARECROW-like, VICILIN 7S, LEAFY COTYLEDON 1, and REVERSIBLE GLYCOSYLATED POLYPEPTIDE 1, were analyzed through semi-quantitative RT-PCR during embryo development and germination. All the eight were found to be differentially expressed in the various developmental stages of zygotic embryos, seeds and seedling tissues. To our knowledge, this is the first report on embryogenesis-regulating gene expression in members of the Araucariaceae family, as well as in plants with recalcitrant seeds

ESTABELECIMENTO IN VITRO DE ÁPICES CAULINARES DE SUMAÚMA (Ceiba pentandra L. Gaertn)

A sumaúma (Ceiba pentandra L. Gaertn) é uma espécie arbórea de grande importância para o setor florestal da Região Norte do Brasil. Apesar disso, ainda não existem relatos sobre o cultivo in vitro desta espécie. O presente trabalho teve como objetivo estabelecer um protocolo eficiente para assepsia e cultivo in vitro de ápices caulinares de sumaúma. Para isto foram avaliados os efeitos de diferentes agentes químicos (NaOCl, HgCl2 e AgNO3) para assepsia e estabelecimento in vitro de ápices caulinares. Diferentes tipos e concentrações de citocininas (BAP, CIN e TDZ) foram testados durante o enraizamento, alongamento e indução de brotos. Apesar das dificuldades usualmente associadas ao estabelecimento e cultivo in vitro de espécies arbóreas, os resultados obtidos não evidenciaram problemas na manipulação in vitro de sumaúma.The kapok tree (Ceiba pentandra L. Gaertn) is a forest species with high importance to the forest sector in North of Brazil. Despite of that, there are no reports about the in vitro culture of this species. The aim of this work was the development of a reliable protocol for disinfestation and in vitro culture of apical shoots of kapok tree. There were tested different chemical compounds (NaOCl, HgCl2 e AgNO3) for apical shoot disinfestation and in vitro establishment. Different types and concentrations of citokinins (BAP, CIN e TDZ) were tested during apical shoot rooting, elongation, and bud induction. Although forest species are normally considered difficult to establish and cultivated in vitro, the kapok tree did not show problems during in vitro manipulation

Biochemical and morphological changes during the growth kinetics of Araucaria angustifolia suspension cultures

Embryogenic cultures of Araucaria angustifolia were established in a BM liquid medium supplemented with 2 µM 2,4-dichlorophenoxyacetic acid, 1 µM 6-benzylaminopurine and 1 µM kinetin (BM2) and in a BM medium free of growth regulators (BM0). During 42 days in culture, the cell growth pattern of both cultures was similar. The pH of the culture medium of both BM0 and BM2 underwent progressive reduction during culture time. For both the embryogenic cultures a preferential uptake of glucose in the late stages of cell growth kinetics was observed. The extracellular protein content was similar for both the embryogenic cultures. Acetocarmine and Evan's blue double stain showed major differences for early somatic embryo organisation, in which only the embryogenic culture grown in a liquid culture medium free of plant growth regulators showed the presence of bipolar somatic pro-embryos.Culturas embriogênicas de Araucaria angustifolia foram estabelecidas em meio de cultura líquido BM suplementado com 2 µM Ácido 2,4 Diclorofenoxiacético, 1 µM 6-Benzilaminopurina e 1 µM Cinetina (BM2) e em meio BM isento de reguladores de crescimento (BM0). Durante 42 dias de cultivo, o padrão de crescimento celular em ambas as culturas foi similar. O pH do meio de cultura BM0 e BM2 sofreu uma progressiva redução durante o período de cultivo. Em ambas as culturas embriogênicas foram observadas um consumo preferencial de glicose no período final da curva de crescimento celular. O nível de proteínas extracelulares foi similar para ambas as culturas embriogênicas. A dupla coloração com carmin acético e azul de Evans revelou diferenças na organização das linhagens celulares embriogênicas, sendo que a presença de proembriões somáticos bipolares foi apenas evidenciada nas culturas embriogênicas mantidas em meio de cultura líquido sem reguladores de crescimento

Partially acetylated chitosan oligo- and polymers induce an oxidative burst in suspension cultured cells of the gymnosperm Araucaria angustifolia

International audienceSuspension-cultured cells were used to analyze the activation of defense responses in the conifer A. angustifolia, using as an elicitor purified chitosan polymers of different degrees of acetylation (DA 1−69%), chitin oligomers of different degrees of polymerization (DP 3−6), and chitosan oligomer of different DA (0−91%). Suspension cultured cells elicited with chitosan polymers reacted with a rapid and transient generation of H2O2, with chitosans of high DA (60 and 69%) being the most active ones. Chitosan oligomers of high DA (78 and 91%) induced substantial levels of H2O2, but fully acetylated chitin oligomers did not. When cultivated for 24−72 h in the presence of 1−10 μg mL−1 chitosan (DA 69%), cell cultures did not show alterations in the levels of enzymes related to defense responses, suggesting that, in A. angustifolia, the induction of an oxidative burst is not directly coupled to the induction of other defense reactions

Chloroplastid pigment contents and chlorophyll a fluorescence in Amazonian tropical three species

Plants react to changes in light and hydrological conditions in terms of quantity and composition of chloroplastidic pigments, which affects the photosynthetic properties and consequently the accumulation of plant biomass. Thus, the chloroplastidic pigment concentration and chlorophyll a fluorescence of three Amazonian species (Bertholletia excelsa, Carapa guianensis e Dipteryx odorata) were investigated in sun and shade leaves form the tree crown collected during two distinct periods of precipitation (dry and rainy seasons). Pigment contents were determined by spectrophotometry and fluorescence variables were determined using a portable fluorometer. The results demonstrated that the species showed high concentrations of Chl a, Chl b e Chl total during the wet season in relation to the dry season, especially in shade leaves. A higher concentration of carotenoids was found in B. excelsa, when compared with leaves of C. guianensis and D. odorata. In leaves of B. excelsa and D. odorata no significant difference was found in relation to the photochemistry of photosystem II (Fv/Fm) between the wet and dry seasons. In conclusion, the three species react differently to variations in the light and precipitation conditions regarding light capture, aspects that might be considered in the management of forest plantations