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Constancia, Portugal

Pinto G.,University of Aveiro | Silva S.,University of Aveiro | Loureiro J.,University of Coimbra | Costa A.,University of Aveiro | And 4 more authors.
Trees - Structure and Function | Year: 2011

This paper reports the complete process from secondary emblings (SE-derived plants) regeneration to acclimatization of Eucalyptus globulus and describes histocytological changes that occur in leaves from in vitro to ex vitro acclimatization for a 3-month period. After elongation, plants were transferred to pots with sterilized peat:perlite and acclimatized in a phytotron, with progressive reduction of RH and increase of light intensity. Histocytological analyses were performed in fixed material using light microscopy and ultrastructural changes followed by electron microscopy (SEM and TEM). The protocol used allowed the successful acclimatization of the emblings. Plants looked morphologically normal and FCM screening revealed no ploidy or DNA content abnormalities. Histocytological analyses showed significant changes along time, mostly in stomata shape and aperture, starch reserves, chloroplast morphology and mesophyll differentiation. This is the first report concerning emblings acclimatization to ex vitro conditions in Eucalyptus. It was clearly demonstrated that during acclimatization emblings suffered profound changes in leaf morphology in order to successfully adapt to ex vitro conditions. © Springer-Verlag.2010.

Pinto G.,University of Aveiro | Silva S.,University of Aveiro | Neves L.,Silvicaima SA Constancia Sul | Araujo C.,Silvicaima SA Constancia Sul | Santos C.,University of Aveiro
Trees - Structure and Function | Year: 2010

We recently described a protocol for Eucalyptus globulus somatic embryogenesis (SE). For its immediate use at industrial levels, some stages of the process require better control. In particular, SE germination rates are variable, decreasing SE efficacy. As reserves may play a central role in embryogenic processes, we followed histocytological changes and reserve fluctuations, during SE. For SE induction, explants of mature zygotic embryos were grown on Murashige and Skoog (MS) medium with 3mgl-1 α-naphthalene acetic acid and later transferred to MS without growth regulators (MSWH). Samples of zygotic embryo cotyledons (explants), of globular and dicotyledonar somatic embryos, and of embling leaves were analysed for reserve accumulation and histocytological profiles. Cotyledon cells of zygotic embryos were rich in lipid and protein bodies, having almost no starch. After 3weeks of induction, starch grain density increased in differentiated mesophyll regions, while in meristematic regions their occurrence was diffuse. In globular somatic embryos, starch accumulation increased with time (in amyloplasts), but protein bodies were absent. Cotyledonary somatic embryos had lower density of starch grains and absence of lipid and protein bodies. Embling leaves showed typical histological organisation. This is the first comprehensive study on histological and cytological changes during Eucalyptus SE with emphasis in reserve accumulation. With this work we demonstrate that the presently available SE protocol for E. globulus leads to reserve fluctuations during the process. Moreover, the reserves of somatic embryo cotyledons differ from those of their zygotic embryo counterparts, which reinforce the importance of reserves in the embryogenic process and suggests that manipulating external conditions, SE may be optimised giving suitable emblings production for industrial purposes. © 2010 Springer-Verlag.

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