Papers by Maria Amely Zavattieri
Methods and Protocols, Jan 7, 2019
An in vitro plant microshoot culture system composed of two phases; a liquid phase overlaid by a ... more An in vitro plant microshoot culture system composed of two phases; a liquid phase overlaid by a floating solid phase, which is described in detail herein. This system is designed to enable the extraction of natural compounds released/disseminated into the liquid phase during root growth, thus facilitating their processing and biochemical characterization. The solid phase holds the plant afloat and enables the simultaneous culture of a microorganism, yet avoiding its penetration into the liquid phase, where the roots are submerged. Both phases can be independently formulated as required for growth optimization of both organisms. Considering the closed system and known variables described in this patent, applications of the described method include testing with pesticides, herbicides, and other similar products.
Methods and Protocols
Several studies have shown the potential of using Ectomycorrhizal (ECM) fungi in conifer micropro... more Several studies have shown the potential of using Ectomycorrhizal (ECM) fungi in conifer micropropagation to overcome the cessation of adventitious root development. In vitro inoculation promotes the re-growth of the root system induced previously by auxin treatments, facilitating acclimation and diminishing the losses of plants because of a weak root system that is incapable of water and nutrient absorption. During a series of mycorrhization experiments, cryostat and ultrafine cuts were used to study the morpho-histological transformation of the symbiotic roots. To obtain cryostat cuts from pine roots a method frequently used for animal tissue was adopted. Molecular methods allowed fungi identification in all the mycorrhization phases and in the acclimation of derived plants. Mycorrhizal-like-structures derived from in vitro culture and axenic liquid cultures of roots were microscopically analyzed and compare with mycorrhizal roots.
Plant Growth Regulation, 65, Apr 1, 2011
Juniperus navicularis Gand (= J. oxycedrus L. subsp. transtagana Franco) is an endemic dioecious ... more Juniperus navicularis Gand (= J. oxycedrus L. subsp. transtagana Franco) is an endemic dioecious shrub from the Plio-Plistocene transition sands of the west Portuguese coast line. It grows in sole or mixed stands along with Pinus pinea and P. pinaster and, more rarely, with Quercus lusitanica. In order to reduce fire incidence in these Mediterranean mixed stands, forest management practices of mechanical or chemical treatments, have drastically reduced J. navicularis populations. In addition, the fast expansion of touristic areas along with a low seed germination rate further endangers this species. In this study we developed a protocol for micropropagation of adult J. navicularis. Microcuttings were obtained from lateral and apical twigs of both female and male mature plants, and used as explants. Microcuttings with axillary buds were grown on different media and plant growth regulators combinations. Developmental parameters were evaluated to define the best medium and plant growth regulator concentration for shoot bud induction during the multiplication cycles. Olive Medium and Gupta and Durzan medium, both supplemented with 0.45 lM 6-benzyl-amino-purine, were the most favourable of all combinations tested achieving more than 3 new shoots per explant. The highest rooting (60%) was obtained in microshoots cultured in Olive Medium when supplemented with indole-3-butyric acid at 12.3 lM. With this multiplication protocol, it was possible to obtain more than 500 shoots from the initial 20, after three multiplication cycles in 3 months.
Electronic Journal of Biotechnology, 2010
Physiologia Plantarum, 2009
Physiologia plantarum, 2009
The gymnosperm Pinus pinea L. (stone pine) is a typical Mediterranean pine used for nuts and timb... more The gymnosperm Pinus pinea L. (stone pine) is a typical Mediterranean pine used for nuts and timber production, and as an ornamental around the world. Pine genomes are large in comparison to other species. The hypothesis that retrotransposons, such as gymny, made a large contribution to this alteration in genome size was recently confirmed. However, P. pinea is unique in other various aspects. P. pinea demonstrates a different pattern of gymny organization than other Pinus subgenera. Additionally, P. pinea has a highly recalcitrant behaviour in relation to standard conifer protocols for the induction of somatic embryogenesis or rooting. Because such types of cell reprogramming can be explained as a reaction of plant cells to external stress, it is of special interest to study sequence peculiarities in stress-inducible genes, such as the alternative oxidase (AOX). This is the first report containing molecular evidence for the existence of AOX in gymnosperms at the genetic level. P. p...
Stone pine (Pinus pinea L.), like other conifers, forms ectomycorrhizas (ECM), which have benefic... more Stone pine (Pinus pinea L.), like other conifers, forms ectomycorrhizas (ECM), which have beneficial impact on plant growth in natural environments and forest ecosystems. An in vitro co-culture of stone pine microshoots with pure mycelia of isolated ECM sporocarps was used to overcome the root growth cessation not only in vitro but also to improve root development during acclimation phase. Pisolithus arhizus (Scop.) Rauschert and Lactarius deliciosus (L. ex Fr.) S.F. Gray fungi, were collected, pure cultured and used in in vitro co-culture with stone pine microshoots. Samples of P. arhizus and L. deliciosus for the in vitro co-cultures were collected from the pine stands southwest Portugal. The in situ characterization was based on their morphotypes. To confirm the identity of the collected material, ITS amplification was applied using the pure cultures derived from the sporocarps. Additionally, a molecular profile using PCR based genomic fingerprinting comparison was executed with other genera of Basidiomycetes and Ascomycetes. Our results showed the effectiveness of the techniques used to amplify DNA polymorphic sequences, which enhances the characterization of the genetic profile of ECM fungi and also provides an option to verify the fungus identity at any stage of plant mycorrhization.
Congreso Forestal Nacional …, 2005
Revista de Ciências …, 2007
La mejora y conservación de recursos genéticos en especies forestales lleva siglos de retraso con... more La mejora y conservación de recursos genéticos en especies forestales lleva siglos de retraso con respecto a las especies agrícolas. Los recursos forestales se han considerado tradicionalmente como recursos "mineros", en los que primaba la mera extracción dejando exclusivamente a la regeneración natural la labor de sostenibilidad en los montes y dehesas o montados. Hoy en día, el necesario desarrollo del medio rural obliga a la explotación racional de los recursos como medio de garantizar su sostenibilidad. Por ello se está empezando a extender el criterio de que las especies forestales se pueden y deben "cultivar" en determinados espacios.
La mejora y conservación de recursos genéticos en especies forestales lleva siglos de retraso con... more La mejora y conservación de recursos genéticos en especies forestales lleva siglos de retraso con respecto a las especies agrícolas. Los recursos forestales se han considerado tradicionalmente como recursos "mineros", en los que primaba la mera extracción dejando exclusivamente a la regeneración natural la labor de sostenibilidad en los montes y dehesas o montados. Hoy en día, el necesario desarrollo del medio rural obliga a la explotación racional de los recursos como medio de garantizar su sostenibilidad. Por ello se está empezando a extender el criterio de que las especies forestales se pueden y deben "cultivar" en determinados espacios.
… on Acclimatization and …, 2007
Clonal propagation via adventitious bud stimulation from cotyledon explants has been reported bef... more Clonal propagation via adventitious bud stimulation from cotyledon explants has been reported before in stone pine (Pinus pinea L.), however the number of rooted shoots had low frequency and short growth span. Therefore the main objective of the present work was to increase the number and quality of roots per shoot in order to ensure the survival and growth of a great number of plants in the acclimation phase. To achieve this, different combinations of carbon source (sucrose or glucose) at different concentrations, under different environmental conditions (temperature and light), were tested in the induction and expression phases of the adventitious root formation. Shoots of different clones obtained via organogenesis have been employed for the experiments. Observations were made on the rooting percentage, root length and number of roots per shoot. The results showed a general increased of the number of roots per shoot and an earlier root formation when glucose was used as a carbon source. However, there were no differences in the percentage of rooting between the carbons sources tested. The best results were obtained using 0.117 M of glucose and dark treatment combined with 19 ºC during the induction phase of the rhizogenic process. Light and low sugar concentration proved to be beneficial for the expression phase, increasing the root length. A remarkable interclonal difference in the ability to form roots was observed. It was possible to obtain a rooting percentage of more than 75% in several of the tested clones.
secforestales.org
Se determinó la capacidad para obtener embriogénesis somática en embriones cigóticos pertenecient... more Se determinó la capacidad para obtener embriogénesis somática en embriones cigóticos pertenecientes a familias de mediohermanos obtenidas de genotipos selectos. La mejor época de recolección resultó ser entre primeros de Agosto y primeros de Septiembre, a partir del momento en que los embriones alcanzaron el estado de desarrollo cotiledonar temprano. La mayor frecuencia de inducción de proliferaciones con características preembriogénicas (PEMs) se obtuvo en medios que contenían NAA y 2.4D como reguladores del crecimiento. Se obtuvieron PEMs en todas las familias analizadas, no encontrándose diferencias familiares significativas en la frecuencia de iniciación. Se han observado diferentes tipos de proliferaciones, algunas de ellas mostrando la presencia de distintas fases del desarrollo de las PEMs, con embriones somáticos en la etapa inicial de su desarrollo acompañados de células de suspensores. La optimización de las condiciones de cultivo, para lograr la proliferación y diferenciación de los embriones somáticos obtenidos en pino piñonero, nos permitirá utilizar esta vía de regeneración de plantas en los programas de mejora y conservación de recursos genéticos de esta especie en nuestro país.
Plant Physiology and …, 2009
The paper highlights Daucus carota L. as an ideal model to complement plant stress research on Ar... more The paper highlights Daucus carota L. as an ideal model to complement plant stress research on Arabidopsis thaliana L. Recently, alternative oxidase (AOX) is discussed as functional marker candidate for cell reprogramming upon stress. Carrot is the most studied species for cell reprogramming and our current research reveals that it is the only one that has expanded both AOX sub-family genes. We point to recently published, but not discussed results on conserved differences in the vicinity of the most active functional site of AOX1 and AOX2, which indicate the importance of studying AOX sequence polymorphism, structure and functionality. Thus, stress-inducible experimental systems of D. carota are especially appropriate to bring research on stress tolerance a significant step forward.
Citação: Ragonezi, C., Klimaszewska, K., Santos-Silva, C., Martins, MR, Caldeira, AT, Zavattieri,... more Citação: Ragonezi, C., Klimaszewska, K., Santos-Silva, C., Martins, MR, Caldeira, AT, Zavattieri, A. 2011. Pisolithus sp. P1001 18S ribosomal RNA gene, partial sequence; internal transcribed spacer 1, 5.8 S ribosomal RNA gene, and internal transcribed spacer ...
ABSTRACT Stone pine (Pinus pinea L.) is an economically important forest tree in the Mediterranea... more ABSTRACT Stone pine (Pinus pinea L.) is an economically important forest tree in the Mediterranean region and has been the target of breeding and selection through micropropagation mainly for its ecological and ornamental aspects. A crucial step in micropropagation is adventitious rooting of microshoots, which often is highly inefficient in most conifer species including stone pine. Hence, we conducted in vitro co-culture of Pinus pinea microshoots with the ectomycorrhizal fungus Pisolithus arhizus (isolated from natural stands) in order to promote adventitious root growth and plant survival during acclimatization. Significant differences were found in the number of branches, in the number of roots plus branches, in total length of roots, in total length of roots plus branches, in average root length and in the length of the longest root in inoculated plants during in vitro rooting compared with non-inoculated plants. The roots of inoculated plants also grew better in vermiculite and during acclimatization in a mixed substrate compared with roots of control plants resulting in the development of vigorous root system. Overall, mycorrhizal inoculation increased the survival rate of the regenerated pine.
The ICAAM Institute aims to study Mediterranean forest ecosystems in all aspects. Our Plant Breed... more The ICAAM Institute aims to study Mediterranean forest ecosystems in all aspects. Our Plant Breeding and Biotechnology Laboratory of ICAAM has always been involved in biotechnology of Mediterranean woody species, and has developed various in vitro ...
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Papers by Maria Amely Zavattieri