Aplicações da micropropagação na clonagem de Eucalyptus no brasil

Presently, in Brazil, among several other in vitro propagation techniques, micropropagation has increased in scientific and economic interest for forestry . Actually, is the most used technique, due its proven practical applications. The first studies were carried out in the 50's; tissue culture laboratories were established during 1970-1980. This article is a review and a synthesis of information on micropropagation applications in Eucalyptus clonal silviculture in Brazil

MORFOANATOMIA DA RIZOGÊNESE ADVENTÍCIA EM MINIESTACAS DE Eucalyptus grandis x Eucalyptus urophylla

http://dx.doi.org/10.5902/1980509815721The present work aimed to accomplish the histological analysis of the events involved in the adventitious rooting pattern of four Eucalyptus grandis x Eucalyptus urophylla clones. The mini-cuttings were collected from a mini-stump and the rooting experiments were set up under mini-hedge indoor semi-hydroponics by means of intermittent flooding fertirrigation. For histological analyses, the proximal ends of the mini-cuttings were collected after 0, 1, 2, 4, 6, 8, 10, and 12 days after planting them at greenhouse. After fixation with FAA70%, the samples were dehydrated in a graded ethanol series and infiltrated overnight in metracrilate resin, and finally embedded in resin. It was verified the endogenous origin of adventitious root primordia from vascular cambium, and in some cases, the formation and proliferation of calli at the proximal end of the mini-cuttings within 8 to 12 days.http://dx.doi.org/10.5902/1980509815721O presente estudo teve como objetivo principal a caracterização morfoanatômica da rizogênese adventícia de miniestacas de quatro clones de Eucalyptus grandis x Eucalyptus urophylla. As miniestacas foram coletadas de minicepas estabelecidas em minijardim clonal, conduzido em sistema semi-hidropônico em canaletas com fertirrigação. Para a caracterização histológica do processo de rizogênese das miniestacas foram coletadas amostras teciduais aos 0, 1, 2, 4, 6, 8, 10 e 12 dias após o estaqueamento em casa de vegetação climatizada, para enraizamento. As amostras foram fixadas em FAA70%, desidratadas em série etílica e incluídas em metacrilato. Os resultados revelaram a origem endógena de primórdios radiculares a partir do câmbio vascular e, em alguns casos, a proliferação e a formação de uma massa de células desorganizadas (calos) em miniestacas dos clones avaliados, entre 8 e 12 dias de idade

MORPHO-ANATOMIC OF ADVENTITIOUS RHIZOGENESIS IN MINI-CUTTINGS OF Eucalyptus grandis x Eucalyptus urophylla

O presente estudo teve como objetivo principal a caracteriza\ue7\ue3o morfoanat\uf4mica da rizog\ueanese advent\uedcia de miniestacas de quatro clones de Eucalyptus grandis x Eucalyptus urophylla . As miniestacas foram coletadas de minicepas estabelecidas em minijardim clonal, conduzido em sistema semi-hidrop\uf4nico em canaletas com fertirriga\ue7\ue3o. Para a caracteriza\ue7\ue3o histol\uf3gica do processo de rizog\ueanese das miniestacas foram coletadas amostras teciduais aos 0, 1, 2, 4, 6, 8, 10 e 12 dias ap\uf3s o estaqueamento em casa de vegeta\ue7\ue3o climatizada, para enraizamento. As amostras foram fixadas em FAA70%, desidratadas em s\ue9rie et\uedlica e inclu\ueddas em metacrilato. Os resultados revelaram a origem end\uf3gena de prim\uf3rdios radiculares a partir do c\ue2mbio vascular e, em alguns casos, a prolifera\ue7\ue3o e a forma\ue7\ue3o de uma massa de c\ue9lulas desorganizadas (calos) em miniestacas dos clones avaliados, entre 8 e 12 dias de idade.The present work aimed to accomplish the histological analysis of the events involved in the adventitious rooting pattern of four Eucalyptus grandis x Eucalyptus urophylla clones. The mini-cuttings were collected from a mini-stump and the rooting experiments were set up under mini-hedge indoor semi-hydroponics by means of intermittent flooding fertirrigation. For histological analyses, the proximal ends of the minicuttings were collected after 0, 1, 2, 4, 6, 8, 10, and 12 days after planting them at greenhouse. After fixation with FAA70%, the samples were dehydrated in a graded ethanol series and infiltrated overnight in metracrilate resin, and finally embedded in resin. It was verified the endogenous origin of adventitious root primordia from vascular cambium, and in some cases, the formation and proliferation of calli at the proximal end of the mini-cuttings within 8 to 12 days

Organogenesis of the leaf explant of Eucalyptus grandis x E. urophylla clones

O objetivo deste trabalho foi avaliar os efeitos dos reguladores de crescimento TDZ [1-fenil-3-(1,2,3-tiadiazol-5-il)uréia], BAP (6-benzilaminopurina) e ANA (ácido naftalenoacético) no desempenho da propagação in vitro por organogênese de explante foliar de três clones híbridos de Eucalyptus grandis x Eucalyptus urophylla. Houve resposta diferenciada dos clones quanto a intensidade, textura e coloração dos calos, em razão dos tratamentos com os reguladores de crescimento. Os melhores resultados de calejamento dos três genótipos foram observados nos tratamentos com a combinação dos reguladores de crescimento TDZ (0,5 mg L-1) e ANA (0,1 mg L-1), obtendo-se 100% de calejamento no explante foliar. Os piores resultados de calejamento foram observados nos tratamentos com a combinação dos reguladores de crescimento BAP (0,1 mg L-1) e ANA (0,1 mg L-1). Em relação à regeneração, a melhor resposta foi obtida com 1,0 mg L-1 BAP em que 8% dos calos formados a partir de explantes foliares regeneraram gemas, com número médio destas formadas por calo igual a 4,2.The aim of this work was to evaluate the effects of growth regulators TDZ [1-phenil-3-(1,2,3-thiadiazol-5-yl)urea], BAP (6-benzilaminopurine) e NAA (Naphthalene acetic acid) on the in vitro propagation by organogenesis from foliar explants of Eucalyptus grandis x E. urophylla. Depending on the clone used, there were singular responses to growth regulators treatment regarding callusing intensity, texture and color. The best results of the three genotypes used were observed with the TDZ (0.5 mg L-1) and NAA (0.1 mg L-1) treatment, where 100% of the foliar explants presented callus. The worst results were observed with the BAP (0.1 mg L-1) and NAA (0.1 mg L-1) treatment. Subsequently, considering the regeneration process, the best response was achieved with 1.0 mg L-1 BAP, in which 8% of the calli regenerated buds, with an average of 4.2 buds per explant

Resgate vegetativo por alporquia de genótipos adultos de urucum (Bixa orellana L.)

This study aimed to evaluate the usefulness of an air layering technique for vegetative rescue of annatto (Bixa orellana L.) genotypes and obtainment of propagules for vegetative propagation. Ten 12 year-old annatto stock plants, whose genotypes are derived from artificial crossing between “Hairy-green capsule” X “Smooth-red capsule” were used in this study. Air layering adventitious roots were induced in branches (1-2 cm diameter) using as substrate a mixture of vermiculite and moss. The effects of (1) the type of girdling (total or partial), 1 cm in length, (2) IBA (indole-3-butyric acid at 0 and 4.92 mM) solution in paper filter and (3) the covering with either transparent or dark plastic films and tencel were evaluated. The technique of air layering provided rooted branches for the 10 genotypes evaluated, with efficiency ranging from 20 to 100%, and a genotype effect was observed on rooting frequency. Under greenhouse conditions, the survival of layers derived from totally girdled stems treated with IBA and protected with transparent plastic was 100%. Established layers provided vigorous branching from which shoot apexes or nodal segment explants were obtained for in vitro establishment and propagation of this species.Este trabalho teve por objetivos avaliar a técnica de alporquia visando ao resgate vegetativo de genótipos de urucum (Bixa orellana L.) e a obtenção de plantas fornecedoras de propágulos para processos de propagação clonal. Foram utilizadas dez plantas matrizes de urucum, com 12 anos de idade, obtidas partindo do cruzamento artificial entre os genótipos "Fruto Verde Piloso" X "Fruto Vermelho Liso". Os alporques foram realizados em ramos de 1 a 2 cm de diâmetro, utilizando-se como substrato uma mistura de vermiculita e musgo. Foi avaliado o efeito (1) do tipo de anelamento da casca dos ramos (total ou parcial), com 1 cm de comprimento, (2) do AIB (ácido indol-3-butírico a 0 e 4,92 mM) aplicado em papel filtro e (3) do tipo de proteção dos alporques (filmes plásticos transparente ou preto ou tecido tencel), no enraizamento de alporque dos dez genótipos. A técnica de alporquia proporcionou o enraizamento de ramos dos dez genótipos avaliados, com eficiência variável de 20 a 100%, havendo efeito do genótipo sobre a frequência de enraizamento. A sobrevivência desses alporques foi de 100% após o plantio quando estes foram produzidos com anelamento total, tratados com AIB e protegidos com plástico transparente. Em casa de vegetação os alporques desenvolveram ramos partindo da brotação de gemas caulinares constituindo estoques de explantes apropriados para serem utilizados como estacas ou como fontes de segmentos nodais para a propagação in vitro desta espécie

Vegetative rescue of adult genotypes of annatto ( Bixa orellana L.) by air layering

Este trabalho teve por objetivos avaliar a t\ue9cnica de alporquia visando ao resgate vegetativo de gen\uf3tipos de urucum ( Bixa orellana L.) e a obten\ue7\ue3o de plantas fornecedoras de prop\ue1gulos para processos de propaga\ue7\ue3o clonal. Foram utilizadas dez plantas matrizes de urucum, com 12 anos de idade, obtidas partindo do cruzamento artificial entre os gen\uf3tipos "Fruto Verde Piloso" X "Fruto Vermelho Liso". Os alporques foram realizados em ramos de 1 a 2 cm de di\ue2metro, utilizando-se como substrato uma mistura de vermiculita e musgo. Foi avaliado o efeito (1) do tipo de anelamento da casca dos ramos (total ou parcial), com 1 cm de comprimento, (2) do AIB (\ue1cido indol-3-but\uedrico a 0 e 4,92 mM) aplicado em papel filtro e (3) do tipo de prote\ue7\ue3o dos alporques (filmes pl\ue1sticos transparente ou preto ou tecido tencel), no enraizamento de alporque dos dez gen\uf3tipos. A t\ue9cnica de alporquia proporcionou o enraizamento de ramos dos dez gen\uf3tipos avaliados, com efici\ueancia vari\ue1vel de 20 a 100%, havendo efeito do gen\uf3tipo sobre a frequ\ueancia de enraizamento. A sobreviv\ueancia desses alporques foi de 100% ap\uf3s o plantio quando estes foram produzidos com anelamento total, tratados com AIB e protegidos com pl\ue1stico transparente. Em casa de vegeta\ue7\ue3o os alporques desenvolveram ramos partindo da brota\ue7\ue3o de gemas caulinares constituindo estoques de explantes apropriados para serem utilizados como estacas ou como fontes de segmentos nodais para a propaga\ue7\ue3o in vitro desta esp\ue9cie.This study aimed to evaluate the usefulness of an air layering technique for vegetative rescue of annatto ( Bixa orellana L.) genotypes and obtainment of propagules for vegetative propagation. Ten 12 year-old annatto stock plants, whose genotypes are derived from artificial crossing between "Hairy-green capsule" X "Smooth-red capsule" were used in this study. Air layering adventitious roots were induced in branches (1-2 cm diameter) using as substrate a mixture of vermiculite and moss. The effects of (1) the type of girdling (total or partial), 1 cm in length, (2) IBA (indole-3-butyric acid at 0 and 4.92 mM) solution in paper filter and (3) the covering with either transparent or dark plastic films and tencel were evaluated. The technique of air layering provided rooted branches for the 10 genotypes evaluated, with efficiency ranging from 20 to 100%, and a genotype effect was observed on rooting frequency. Under greenhouse conditions, the survival of layers derived from totally girdled stems treated with IBA and protected with transparent plastic was 100%. Established layers provided vigorous branching from which shoot apexes or nodal segment explants were obtained for in vitro establishment and propagation of this species

In vitro grafting in the propagation of Eucalyptus urophylla and E. grandis clones

O objetivo deste trabalho foi avaliar a eficiência da enxertia in vitro na propagação de dois clones híbridos de Eucalyptus urophylla e E. grandis. Foram utilizados porta-enxertos juvenis obtidos de sementes de E. grandis e E. urophylla germinadas in vitro. As plantas enxertadas apresentaram pegamento de até 93% aos 50 dias de idade para as combinações de enxerto e porta-enxerto. Quanto ao crescimento em altura, verificaram-se melhores resultados em relação ao porta-enxerto E. urophylla. A análise histológica das plantas enxertadas comprovou a eficiência do processo de cicatrização do calo formado na região de conexão, seguida da reconstituição vascular.The objective of this work was to evaluate the efficiency of the in vitro grafting in the propagation of two Eucalyptus urophylla and E. grandis hybrid clones. Juvenile stocks obtained from seeds of E. grandis and E. urophylla germinated in vitro were used. The grafted plants showed success up to 93% at 50 days for the scion and stock combinations. The highest final plant height values were observed when the stock was E. urophylla. Histologic analysis showed that cicatrized callus in the connection region was successfully followed by vascular recomposition in different combinations among scion with rootstocks

MULTIPLICAÇÃO E ALONGAMENTO IN VITRO DE CLONES HÍBRIDOS DE Eucalyptus globulus

http://dx.doi.org/10.5902/1980509821524Eucalyptus globulus hybrids represent an excellent alternative to the pulp and paper sector, because of gains in wood quality for pulp. However, these hybrids have shown recalcitrance to adventitious rooting. Thus, micropropagation is considered the technique for rejuvenating adult genetic materials, allowing its clonal propagation. This study evaluated the in vitro culture of three clones of Eucalyptus grandis x Eucalyptus globulus and three clones of Eucalyptus urophylla x Eucalyptus globulus in relation to in vitro multiplication on MS medium supplemented with 0.5 mgL-1 BAP and 0,01 mgL-1 NAA. The effect of IBA concentrations (0.25; 0.50; 0.75 and 1.0 mg L-1) and MS and JADS culture media was evaluated on in vitro elongation. The clones differed during in vitro multiplication of shoots and they had an average multiplication rate of 3.0 shoot clusters in each subculture. The responses of the clones to different concentrations of IBA, as well as to MS and JADS culture media differed for elongation in vitro. The range between 0.40 and 0.80 mg L-1 AIB provided the highest number and length of in vitro elongated shoots and the best vigor of shoots.http://dx.doi.org/10.5902/198050982524Os híbridos de Eucalyptus globulus representam uma excelente alternativa para o setor de celulose e papel, em razão dos ganhos em qualidade da madeira para a fabricação de celulose. Entretanto, estes híbridos têm apresentado recalcitrância ao enraizamento adventício. Assim, a micropropagação é apontada como a técnica para o rejuvenescimento desses híbridos adultos, viabilizando a propagação clonal dos mesmos. O presente trabalho avaliou o cultivo in vitro de três clones de Eucalyptus grandis x Eucalyptus globulus e de três clones de Eucalyptus urophylla x Eucalyptus globulus, em relação à multiplicação in vitro, no meio MS suplementado com 0,5 mg L-1 de BAP e 0,01 mg L-1 de ANA, bem como o efeito das concentrações de 0,25; 0,50; 0,75 e 1,0 mg L-1 de AIB e dos meios de cultura MS e JADS no alongamento in vitro das brotações. Os clones diferiram quanto à multiplicação in vitro das brotações e apresentaram uma taxa de multiplicação média dos clones de 3,0 tufos de brotações em cada subcultivo, ao longo dos 25subcultivos realizados. No alongamento in vitro, os clones diferiram quanto às concentrações de AIB adequadas para provocar o alongamento, bem como em relação aos meios de cultura MS e JADS. O intervalo médio entre 0,40 e 0,80 mg L-1 de AIB proporcionou o maior número e comprimento das brotações alongadas in vitro e com maior vigor.

IN VITRO MULTIPLICATION AND ELONGATION OF HYBRID CLONES OF Eucalyptus globulus

Os h\uedbridos de Eucalyptus globulus representam uma excelente alternativa para o setor de celulose e papel, em raz\ue3o dos ganhos em qualidade da madeira para a fabrica\ue7\ue3o de celulose. Entretanto, estes h\uedbridos t\ueam apresentado recalcitr\ue2ncia ao enraizamento advent\uedcio. Assim, a micropropaga\ue7\ue3o \ue9 apontada como a t\ue9cnica para o rejuvenescimento desses h\uedbridos adultos, viabilizando a propaga\ue7\ue3o clonal dos mesmos. O presente trabalho avaliou o cultivo in vitro de tr\ueas clones de Eucalyptus grandis x Eucalyptus globulus e de tr\ueas clones de Eucalyptus urophylla x Eucalyptus globulus, em rela\ue7\ue3o \ue0 multiplica\ue7\ue3o in vitro, no meio MS suplementado com 0,5 mg L-1 de BAP e 0,01 mg L-1 de ANA, bem como o efeito das concentra\ue7\uf5es de 0,25; 0,50; 0,75 e 1,0 mg L-1 de AIB e dos meios de cultura MS e JADS no alongamento in vitro das brota\ue7\uf5es. Os clones diferiram quanto \ue0 multiplica\ue7\ue3o in vitro das brota\ue7\uf5es e apresentaram uma taxa de multiplica\ue7\ue3o m\ue9dia dos clones de 3,0 tufos de brota\ue7\uf5es em cada subcultivo, ao longo dos 25 subcultivos realizados. No alongamento in vitro, os clones diferiram quanto \ue0s concentra\ue7\uf5es de AIB adequadas para provocar o alongamento, bem como em rela\ue7\ue3o aos meios de cultura MS e JADS. O intervalo m\ue9dio entre 0,40 e 0,80 mg L-1 de AIB proporcionou o maior n\ufamero e comprimento das brota\ue7\uf5es alongadas in vitro e com maior vigor.Eucalyptus globulus hybrids represent an excellent alternative to the pulp and paper sector, because of gains in wood quality for pulp. However, these hybrids have shown recalcitrance to adventitious rooting. Thus, micropropagation is considered the technique for rejuvenating adult genetic materials, allowing its clonal propagation. This study evaluated the in vitro culture of three clones of Eucalyptus grandis x Eucalyptus globulus and three clones of Eucalyptus urophylla x Eucalyptus globulus in relation to in vitro multiplication on MS medium supplemented with 0.5 mgL-1 BAP and 0,01 mgL-1 NAA. The effect of IBA concentrations (0.25; 0.50; 0.75 and 1.0 mg L-1) and MS and JADS culture media was evaluated on in vitro elongation. The clones differed during in vitro multiplication of shoots and they had an average multiplication rate of 3.0 shoot clusters in each subculture. The responses of the clones to different concentrations of IBA, as well as to MS and JADS culture media differed for elongation in vitro. The range between 0.40 and 0.80 mg L-1 AIB provided the highest number and length of in vitro elongated shoots and the best vigor of shoots

Involvement of glutathione metabolism in Eichhornia crassipes tolerance to arsenic

Aquatic macrophytes are potential useful for phytoremediation programs in environments contaminated by arsenic (As). Biochemical and physiological modification analyses in different plant parts are important to understand As tolerance mechanisms. 2. The objective was to evaluate glutathione metabolism in leaves and roots of Eichhornia crassipes (Mart.) Solms plants subjected to As. Specimens of Eichhornia crassipes plants were cultured for three days in Clark's nutrient solution containing 7 μM of As. The enzymes ATP sulfurylase (ATPS), glutathione reductase (GR), glutathione peroxidase (GSH‐Px), glutathione sulfotransferase (GST) and γ‐glutamylcysteine (γ‐ECS) synthetase activity, the glutathione contents, total proteic and non‐proteic thiols were evaluated. 3. The ATPS activity increased in roots. The GR activity in leaves and GSH‐Px in roots were lower. GST activity was higher in roots and lower in leaves and γ‐ECS activity was higher in this plant's leaves. Glutathione levels were lower, total thiols levels were higher and non proteic levels presented no change in E. crassipes leaves and roots. Exposure to As increased the enzyme activity involved with sulfur metabolism, such as ATPS. Higher GR activity and lower GSH‐Px indicates a greater glutathione conjugation to As, due to the greater GSH availability. The greater GST activity indicates its participation in As detoxification and accumulation, through As GSH conjugation. Changes in the glutathione and thiol levels suggest high phytochelatin synthesis. 4. In conclusion, the increment in ATPS, GR, GST and γ‐ECS activity indicates that these enzymes are involved in GSH metabolism and are part of the E. crassipes As detoxification mechanism