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In vitro plantlet regeneration from mature zygotic embryos of Pinus pinea L.: overcoming the rooting problems
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Authors
Abstract(s)
Na natureza, as plantas e os microrganismos estabelecem associações de várias
ordens. Nas culturas in vitro de plantas, mesmo as associações favoráveis com
microrganismos foram, durante muitos anos, consideradas como contaminantes. Só
mais tarde, as vantagens da inoculação in vitro (co-cultura) foram demonstradas e as
técnicas de biotização (bacterização ou micorrização) usadas com o objetivo de
melhorar as condições de crescimento in vitro.
As dificuldades do enraizamento in vitro de uma das espécies mais importantes da
floresta mediterrânica portuguesa, Pinus pinea L., conduziu à escolha deste sistema
biológico, como matéria de estudo para a tese. Neste estudo, foram utilizados fungos
ectomicorrízicos para otimizar a fase de enraizamento de plantas de Pinus pinea L.
micropropagadas via organogénese. A introdução de ectomicorrízas no processo de
micropropagação reativou o crescimento das raízes e induziu a melhoria dos vários
parâmetros do sistema radicular adventício conduzindo a uma menor perda de plantas
durante a aclimatização. Com efeito, a micorrização melhorou a funcionalidade das
raízes, facilitando a absorção de nutrientes e de água.
Neste trabalho, efetuou-se também uma extensiva caracterização morfológica e
molecular das ectomicorrízas associadas a P. pinea. Das várias co-culturas testadas,
selecionou-se a interação Pisolithus arhizus/P. pinea para estudar os sinais bioquímicos
pré-simbióticos estabelecidos durante as etapas iniciais da co-cultura in vitro. Para
possibilitar este estudo foi desenvolvido um novo sistema de co-cultura, o qual já está
patenteado. Os resultados indicam que a presença de compostos fenólicos,
nomeadamente o ácido o-coumarico, poderão ser importantes mediadores na interação
fungo/planta.
In nature, plants and microorganisms establish symbiotic associations of various orders. However, for many years such associations were deemed unnecessary in in vitro cultures because the culture medium provides ample amounts of nutrients and plant growth regulators to a growing plant. Only recently, the benefits of biotization (bacterization or mycorrhization) of plants regenerated in vitro were demonstrated by improvements in their growth and vigor. Pinus pinea L., which is of one of the most important species of Portuguese Mediterranean forests, can be regenerated in vitro from embryo cotyledons but the growth of adventitious roots induced in shoots ceases shortly after their formation. Overcoming this particular biological impediment was the study subject of the thesis. In this study, ectomycorrhizal fungi were used to improve adventitious rooting of Pinus pinea L. plants micropropagated through organogenesis. The introduction of ectomycorrhizae during the micropropagation process reactivated the root growth and improved several root characteristics leading to a reduced loss of plants during acclimatization. In fact, the mycorrhization enhanced root functionality facilitating the absorption of nutrients and water. In this work, an extensive characterization of morphological and molecular ectomycorrhizae associated with P. pinea was also undertaken. Of the several fungus species tested, the interaction of Pisolithus arhizus/P. pinea was selected for studying the pre-symbiotic biochemical signals established during the initial stages of co-culture in vitro. To facilitate this study, a novel co-culture system was developed which has been patented. The results indicate that the phenolic compounds, in particular the o coumaric acid ester might be important mediators in the interaction between the fungus and stone pine.
In nature, plants and microorganisms establish symbiotic associations of various orders. However, for many years such associations were deemed unnecessary in in vitro cultures because the culture medium provides ample amounts of nutrients and plant growth regulators to a growing plant. Only recently, the benefits of biotization (bacterization or mycorrhization) of plants regenerated in vitro were demonstrated by improvements in their growth and vigor. Pinus pinea L., which is of one of the most important species of Portuguese Mediterranean forests, can be regenerated in vitro from embryo cotyledons but the growth of adventitious roots induced in shoots ceases shortly after their formation. Overcoming this particular biological impediment was the study subject of the thesis. In this study, ectomycorrhizal fungi were used to improve adventitious rooting of Pinus pinea L. plants micropropagated through organogenesis. The introduction of ectomycorrhizae during the micropropagation process reactivated the root growth and improved several root characteristics leading to a reduced loss of plants during acclimatization. In fact, the mycorrhization enhanced root functionality facilitating the absorption of nutrients and water. In this work, an extensive characterization of morphological and molecular ectomycorrhizae associated with P. pinea was also undertaken. Of the several fungus species tested, the interaction of Pisolithus arhizus/P. pinea was selected for studying the pre-symbiotic biochemical signals established during the initial stages of co-culture in vitro. To facilitate this study, a novel co-culture system was developed which has been patented. The results indicate that the phenolic compounds, in particular the o coumaric acid ester might be important mediators in the interaction between the fungus and stone pine.
Description
Keywords
Adventitious rooting Biotization Ectomycorrhiza in vitro mycorrhization Micropropagation Pinus pinea L. Biology . Escola Superior de Tecnologias e Gestão