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The auxins, including indole-3-acetic acid (IAA), indole-3-butyric acid (IBA) and naphthaleneacetic acid (NAA), promotes the formation of adventitious root in hypocotyls of soybean (Glycine max). IBA significantly promotes the formation of adventitious roots more than IAA and NAA . The activity of anionic pI 3.7 peroxidase (POX) and cationic pI 8.5 POX were inhibited by exogenous auxins during the induction of adventitious root on day 2. Besides, the activity of pI 5.3 POX was enhanced by IBA during the initiation stage on day 4. The increase of the activity of pI 5.3 POX was accompanied by the increase of H2O2 levels. In the previous researches, it shows that the promoter of pI 8.5 POX gene contains both ARF/AuxRE and CATATGGMSAUR motifs that are responded to auxins. In this studies, the pI 5.3 POX gene, which responses to auxins a day or two later, contains only ARF/AuxRE motif. The regulation of pI 5.3 POX gene is probably initiation phase-dependent. The results suggest that anionic pI 5.3 POX produces significant amount of H2O2 through the binding of auxin to POX and mediate the auxin signaling pathway leading to plant growth .

The objectives of this study were to develop an in vitro direct adventitious bud induction and an organogenic callus induction and shoot regeneration system via floral organ segments culture for bromeliads, moreover, explore the effect of auxin on plantlet elongation of Guzmania. And further, apply the above micropropagation system to physical and chemical methods to induce somaclonal variances of bromeliad plantlets in vitro for mutation breeding. The explant sources of bromeliads and the components of culture medium were studied to develop a micropropagation system for bromeliads. The results indicated that the 1/3MS basal medium supplemented with a combination of 1.0 mg l-1 BA + 0.5-1.0 mg l-1 NAA, or a combination of 3.0 mg l-1 BA + 0.5 mg l-1 NAA, showed the highest frequency of direct initiation of adventitious buds derived from shoot apex and lateral bud explants of Aechmea fulgens var. fulgens and Guzmania 'Focus'. The best results of adventitious buds induction of the both species were found in the lower lateral bud explants, at 47.5% and 35%, respectively. In addition, the adventitious buds began to form on day 16 after the G. 'Focus' decapitated plantlets had been cultured in medium supplemented with 3.0 mg l-1 BA + 0.5 mg l-1 NAA. However, this phenomenon did not occur in case of undecapitated explants, where only protruding nodules appeared. Petal- and ovary-derived calli of A. fasciata and G. 'Hilda' were induced on 1/2MS basal medium supplemented with 1.0-1.5 mg l-1 2,4-D in combination with 1.0 or 0.5 mg l-1 NAA. Organogenic calli were cultured on medium with 1.0 mg l-1 NAA and 0.5 mg l-1 TDZ could be induced to differentiate and regenerate the adventitious buds. Furthermore, the number of adventitious buds proliferating at the base of the plantlets derived from G. 'Hilda' floral organs, cultured in media with different concentrations of IAA, IBA, NAA, and 8-azaadenine, was only 1-2 adventitious buds individually. This result shows that auxin can indeed suppress cytokinin-effects. The influence on plantlet elongation was greatest in the treatments using 0.5 mg l-1 NAA and 1.0 mg l-1 NAA. After 4 months culture, plantlets grew to 5.73 and 5.62 cm in height, that was 2.22 and 1.95 cm higher than the control, respectively. Plantlets of A. fasciata hardened under the middle (50 £gmol m-2s-1) light intensity condition had a higher survival rate, 95%, than that hardened at a low light intensity (1 £gmol m-2s-1; 17.5%). The maximum number of newly developing roots, up to 4.15 per shoot, was also observed at the same light intensity treatment. During transplantation, plantlets growing in coir fiber showed the best results in terms of plant growth within 6 months ex vitro culture. The average length of the plantlets was 22.0 cm, and an average of 19.3 leaves per plantlet was achieved. When calli of G. 'Hilda' treated by sodium azide, the survival rate was 0%. The survival rate of decapitated plantlet explants treated with 0.5 mM sodium azide for 60 minutes was 51.3%, about half-lethal dose. In addition to the survival rates of decapitated plantlet explants of A. fasciata, G. 'Hilda', G. 'Cherry', G. 'Luna' and G. 'Focus' irradiated by £^-ray showed 74.2-100% with the exception of the G. 'Focus' irradiated by 15 Gy, which dropped to 45.0%. At present, mutant plantlets showed a great deal of chimeras in leaf and were transplanted to potting media.

Synthetic auxin, naphthaleneacetic acid ( NAA ), promoted rooting in soybean ( Glycine max ) hypocotyls. The activities of anionic peroxidases (PODs, pI 3.5 and pI 3.7 ) and cationic PODs ( pI 8.3, pI 9.1 and pI 9.4 ) in soybean hypocotyls were significantly inhibited by exogenous NAA during the induction of adventitious roots. In addition, the reduction in the activity of PODs ( pI 4.4, pI 8.3 and pI 9.4 ) were correlated with a reduction of their corresponding transcription activity. The PODs patterns and transcription in the NAA-treated hypocotyls demonstrated the specific effect of NAA on POD ioszymes, indicating that NAA might suppress PODs genes expression during the adventitious root formation. The decline of PODs activity in the NAA-treated hypocotyls was accompanied by the decrease of lignin contents. We suggest that both of anionic and cationic PODs might be in part responsible for lignin synthesis in soybean hypocotyls.

The objective of the study was to investigate the optimum conditions for in vitro regeneration of common juniper (Juniperus communis L.) by using indirect organogenesis approach. Throughout the study / callus induction, organogenesis, improved organogenesis and root induction experiments were performed sequentially. It was found that explant position, genotype, gender, treatments and sampling time had significant effects on callus induction rate in common juniper. The results of treatments indicated that IBA (indole-3-butyric acid) at concentration range 0.5-4.0 mg/l combined with MS medium supplemented with 0.1 mg/l BAP (benzylaminopurine), 3 % sucrose and 0.7% agar was the best one among the treatments to induce callus formation from common juniper explants collected as Spring buds. Also, a two-month culture was adequate period for the callus induction of common juniper regardless of position, before transferring the explants into organogenesis media. After a two-month culture in callus induction media, explants were transferred to organogenesis treatments in order to investigate adventitious bud development from callus tissues. There were significant differences among genotypes, treatments and explant-sampling times in initiation of organ development in common juniper. Additionally, it was found that excluding the auxin components while maintaining 1.0-2.0 mg/l BAP concentration in culture media, as refreshing after a month, stimulated the formation and development of adventitious buds and shoots. Among the treatments tested, it was found that 1.0 mg/l BAP plus 0.5 mg/l 2,4-D was the optimum culture media with adventitious bud formation capacity of 37.5% was though ageing of callus significantly affected the frequency of adventitious bud formation. Finally, rooting experiments were performed to investigate rooting efficiency of adventitious shoots. In the adventitious rooting experiments, no rooting was observed in any of the treatments used with common juniper explants. Although whole plantlet development from callus tissues could not be achieved as indirect organogenesis, the results of the study could aid to future studies dealing in vitro regeneration and production of secondary chemicals from common juniper.

A família Bromeliaceae apresenta importância ecológica e econômica, desta forma, o desenvolvimento de protocolos para a micropropagação de espécies dessa família, faz-se necessário, a fim de suprir sua demanda comercial e mesmo ecológica. A escolha do meio de cultura e do explante utilizado durante a micropropagação são fundamentais para um protocolo eficaz. Nesse contexto, o objetivo deste estudo foi avaliar as diferenças quantitativas e qualitativas no desenvolvimento de explantes de Neoregelia sp em meios de cultura e monitorar as vias morfogênicas dos propágulos obtidos em explantes foliares. Para tanto, brotos de microcepas e explantes foliares procedentes de um micro jardim clonal, foram transferidos para os meios de cultura de multiplicação MS, ½ MS e WPM, todos suplementados com 0,050 mg.L-1 ANA e 0,50 mg.L-1 de BAP, onde foram mantidos por 120 dias e submetidos a diversas análises morfofisiológicas. Paralelamente, explantes foliares foram mantidos em meio de cultura MS de multiplicação para o monitoramento das vias morfogênicas durante os processos regenerativos. Para os experimentos com brotos de microcepas verificou-se que o meio de cultura MS proporcionou a melhor taxa de multiplicação, maior crescimento dos brotos, obtendo os valores mais elevados de peso de matéria fresca e seca, além disso, apresentaram maior acúmulo de nitrogênio total e proteico. No entanto, os meios de cultura ½ MS e WPM promoveram uma taxa de multiplicação semelhante a do MS, mas com brotos menores e menos vigorosos, porém, mais homogêneos, com isso, na dependência do objetivo do cultivo in vitro, não deve ser desconsiderada a possibilidade de utilização dos meios de cultura ½ MS e WPM. Os explantes foliares não se desenvolveram bem no meio de cultura WPM, não havendo diferença entre os meios MS e ½ MS, visto que ambos apresentaram resultados satisfatórios. As análises histológicas e histoquímicas identificaram células parenquimáticas, que atuam como células-tronco, manifestando capacidade morfogênica para toti ou pluripotência, dando origem respectivamente a embriões somáticos e gemas adventícias, em resposta aos estímulos in vitro. / The Bromeliaceae family has an ecological and economic importance, therefore, the protocols development for micropropagation of species of this family becomes necessary in order to meet its business and even its ecological demand. The choice of culture medium and the explant used during micropropagation are essential for an effective protocol. Thus, the aim of this study was to evaluate the quantitative and qualitative differences in the explants development of Neoregelia sp in the culture media and monitor the morphogenetic pathways of obtained propagules from leaf explants. Consequently, shoots and leaf explants coming from microcloning garden were transferred to the MS, ½ MS and WPM multiplication culture media, all supplemented with 0.050 mg.L-1 NAA and 0.50 mg.L-1 BAP, where they were held for 120 days and submitted to morphological and physiological analysis. Therefore, leaf explants were kept on MS-medium multiplication for monitoring morphogenetic pathways during the regenerative processes. Furthermore, MS medium showed the best multiplication rate for the sprouts of the microstumps, increased growth of shoots, obtaining the highest values of fresh and dry matter weight, and also showed higher accumulation of total nitrogen and protein. However, the ½ MS and WPM culture media promoted a similar multiplication rate to the MS medium, with the development of the smaller and less vigorous shoots, but with greater homogeneity. This way, depending on the purpose of in vitro culture, their use in the micropropagation for this species should not be disregarded. The leaf explants are not well developed in WPM medium, and don\'t had significant difference between the MS and ½ MS culture medium, as both showed satisfactory results. The histological and histochemical analysis identified the presence of the parenchymatic cells, which act as stem cells, expressing morphogenic ability for toti or pluripotency, leading respectively to somatic embryogenesis or adventitious organogenesis in response to in vitro stimuli.

O Pinus elliotti representa o principal produtor atual de exploração de resina no setor florestal brasileiro. Recentemente, plantios de P. elliottii X P. caribaea vêm se destacando, principalmente, pela sua maior produtividade em biomassa. Além disto, a resina do híbrido apresenta características químicas semelhantes aos seus parentais. Porém, poucas são as informações quanto a estaquia de espécies do gênero Pinus, e se focarmos na propagação de árvores adultas, tal carência é ainda maior, principalmente ao se considerar a dificuldade de rejuvenescimento/revigoramento do material genético. Neste contexto, o presente trabalho teve como objetivo geral avaliar o enraizamento adventício do P. elliottii var. elliottii e P. elliottii X P. caribaea adultos. Para tanto, o trabalho foi dividido em três diferentes estudos. O primeiro (Capítulo 2) avaliou o enraizamento adventício de estacas das matrizes adultas de P. elliottii e P. elliottii X P. caribaea no 5° estágio de enxertia seriada com e sem podas sucessivas, sob ação do AIB, talco com fungicida Cerconil® e terebentina. O segundo (Capítulo 3) foi baseado na avaliação do enraizamento adventício de estacas no 2° estágio de enxertia seriada de uma matriz adulta de P. elliottii var. elliottii submetida a um ano de podas sucessivas, sob ação do AIB, terebentina e paclobutrazol. Por sua vez, o terceiro estudo (Capítulo 4) consistiu na avaliação do enraizamento adventício de braquiblastos e brotos de duas matrizes adulta de P. elliottii e P. elliottii X Pinus caribaea, sob ação do AIB e paclobutrazol. Os resultados permitiram inferir que a sobrevivência e o enraizamento adventício aos diferentes tratamentos variaram de acordo com o genótipo e com as características morfológicas das estacas (grau de lignificação e tipo de estacas, brotos e braquiblastos). De maneira geral, os resultados evidenciaram a atuação das enxertias seriadas e das podas sucessivas na restauração da competência rizogênica das estacas dos brotos das matrizes adultas de P. elliottii e P. elliottii X P. caribaea, promovido pelo rejuvenescimento/ revigoramento parcial das matrizes. Além disto, a estaquia de braquiblastos mostrou ser uma ferramenta promissora para a propagação de espécies do gênero Pinus. / Pinus elliottii is the main current resin extraction producer in the Brazilian forest sector. Recently, plantations of P. elliottii X P. caribaea play a great importance in the production systems mainly by its high biomass productivity. In addition, the hybrid resin has chemical characteristics similar to their parents. However, there is little information on obtaining clones, and considering the propagation of mature trees, this lack of information is even greater, mainly concerning the difficulty to achieve the rejuvenation/ reinvigoration of the genetic material. Based on theses information, the present work was aimed to evaluate the adventitious rooting of mature P. elliottii var. elliottii and P. elliottii X P. caribaea cuttings. Therefore, the work was divided into three basic studies. The first study (Chapter 2) was based on the evaluation of adventitious rooting of mature P. elliottii and P. elliottii X caribaea cuttings of grafts up five sequential grafting with and without pruning, under the action of AIB, talc with fungicide and turpentine. The second (Chapter 3) was based on the evaluation of adventitious rooting of mature P. elliottii cuttings of grafts up two sequential grafting with pruning, under the action of IBA, paclobutrazol and turpentine. The third (Chapter 4) evaluated the adventitious rooting of brachyblast and shoots of mature P. elliottii and P. elliottii X caribaea cuttings, under the action of IBA and paclobutrazol. Results showed that the cutting survival and root growth, under the action of different treatments, varied according to the genotype and morphological characteristics of the material (cutting lignifications and type, brachyblast or shoot). Overall, our results showed the performance of serial grafting and pruning on the restoration of rooting competence in P. elliottii and P. elliottii X P. caribaea stem cuttings, promoted by the rejuvenation of trees. Moreover, the cutting of brachyblast can be a promising tool for the propagation of Pinus species.

Poucas espécies de Eucalyptus apresentam aptidão ao cultivo em regiões de baixas temperaturas e geadas frequentes, limitando seu cultivo em algumas regiões do Brasil. Genótipos de Eucalyptus globulus podem representar opções futuras para plantios florestais, tendo em vista o seu ótimo desempenho silvicultural nessas condições. No entanto, informações quanto à obtenção de mudas clonais são escassas, e se focarmos as espécies recomendadas para plantio em condições subtropicais, tal carência é ainda maior, principalmente em termos endógenos e exógenos de enraizamento de propágulos. Esta espécie se destaca também, pelo grande interesse no setor florestal, sobretudo em relação às características de sua madeira para obtenção de celulose, porém, é considerada recalcitrante ao enraizamento de estacas, especialmente quando envolve material adulto, dificultando o aproveitamento dos benefícios da clonagem. Por essa razão, o presente trabalho teve como objetivo principal avaliar a técnica de micropropagação de clones de E. globulus, visando a reversão à juvenilidade, aumentando os índices de enraizamento e, consequentemente a produção de mudas clonais. Para tanto, os clones (USP 01, USP 24 e USP 33) foram estudados e apresentaram estabelecimento satisfatório, onde avaliou-se: (a) a multiplicação in vitro, testando-se três meios de cultura (WPM, JADS e MS) suplementados com concentrações do regulador de crescimento BAP (0; 0,50 e 1,0 mg L-1) combinado com ANA (0; 0,05 e 0,10 mg L-1); (b) o alongamento in vitro, testando-se o meio de cultura WPM suplementado com concentrações de BAP (0; 0,05 e 0,10 mg L-1) combinadas com ANA (0; 0,1; 0,2 e 0,3 mg L-1); (c) e o enraizamento in vitro, em que brotações alongadas in vitro dos clones foram coletadas e transferidas para meio de cultura WPM, suplementado com 0 e 0,05 mg L-1 de BAP combinado com 0; 0,1; 0,5 e 1,0 mg L-1 de AIB. Os resultados revelaram comportamentos diferenciados dos clones quanto à multiplicação e alongamento in vitro. As taxas de multiplicação in vitro dos clones variaram ao longo dos 10 subcultivos, com uma estabilização a partir do 6º subcultivo para todos os clones avaliados, sendo a maior taxa de multiplicação (número médio de 13 gemas) na concentração de 0,7 mg L-1 de BAP e 0,05 mg L-1 de ANA no meio de cultura WPM. O meio de cultura WPM avaliado no alongamento in vitro, contribuiu favoravelmente para o alongamento das brotações, assim como ANA na concentração de 0,3 mg L-1 combinando com 0,1 mg L-1 de BAP. Portanto, o protocolo desenvolvido mostrou-se eficiente para micropropagação via proliferação de gemas axilares dos clones avaliados, porém, em relação ao enraizamento constatou-se apenas uma raiz emitida na concentração de 0,5 mg L-1 de AIB no clone 01. De forma geral, em razão dos resultados observados, conclui-se que apesar de mostrar-se viável, a micropropagação não promoveu rejuvenescimento significativo dos clones de E. globulus, uma vez que não foi observado enraizamento expressivo das microestacas para as características avaliadas. / Few Eucalyptus species present adaptation for cultivation in regions with low temperatures and frequent frosts, limiting their cultivation in some regions of Brazil. E. globulus genotypes may to represent future options for forest plantations, in view of its optimal silvicultural performance in these conditions. However, information about obtaining clonal seedlings are scarce, and considering the species recommended for planting in subtropical conditions, this lack of information is even greater, mainly when considering the endogenous and exogenous factors for the rooting. This species is distinguished by great interest in forestry, particularly in relation to characteristics of its wood to obtain cellulose, however, it is considered recalcitrant to rooting, especially when it involves adult material, making it difficult to utilize the cloning benefits. Therefore, this study aimed to evaluate the micropropagation technique of E. globulus clones, in order to revert to juvenility, increasing the rate of rooting and consequently the clonal production. For this, the clones (USP 01, USP 24 and USP 33) were studied and showed satisfactory in vitro culture establishment, where it was evaluated: (a) the in vitro multiplication, testing three culture media (WPM, JADS and MS) supplemented with concentrations plant growth regulator BAP (0; 0,50 and 1,0 mg L-1) combined with NAA (0; 0,05 and 0,10 mg L-1); (b) the in vitro elongation, testing the WPM medium culture supplemented with BAP concentrations (0; 0,05 and 0,10 mg L-1) combined with NAA (0; 0,1; 0,2 and 0,3 mg L-1); and (c) in vitro rooting, where elongated shoots in vitro of the clones were collected and transferred to WPM, supplemented with 0 and 0,05 mg L-1 BAP combined with 0; 0,1, 0,5 and 1,0 mg L-1 IBA. The results showed different behaviors of clones as in vitro multiplication and elongation. Rates of clones in vitro multiplication varied over the 10 subcultures, with the stabilization from the sixth subculture for all clones, the largest rate multiplication (average number of 13 buds) in the concentration of 0,7 mg L-1 BAP and 0,05 mg L-1 NAA in WPM. The culture media WPM evaluated in vitro elongation, contributed positively to the shoots elongation, with 0,3 mg L-1 NAA and 0,1 mg L-1 BAP. Therefore, with the micropropagation protocol using axillary buds proliferation of these clones showed be well developed , however, in relation to rooting, only one root was emitted at 0,5 mg L-1 IBA in clone 01. In general, it is possible concluded that despite viable, the micropropagation did not promote significant rejuvenation of E. globulus clones, due to the not expressive rooting for the microcutting evaluated.

Flax is one of the major cash crops in Canada. Approximately seventy percent of Canadian flaxseed was exported to European Union (EU) annually until 2009. In 2009, the EU imposed an import ban on Canadian flaxseed due to the adventitious presence of a GM flax variety - CDC Triffid was identified in Canadian flaxseed exported to the EU. The EUs decision to apply zero tolerance on CDC Triffid flax has been based on its interpretation of the precautionary principle. According to the World Trade Organisations Agreement on the Application of Sanitary and Phytosanitary Measures (SPS), however, precautionary measures are subject to a scientific risk assessment. As the EU did not base its zero tolerance for CDC Triffid flax on any scientific risk assessment, the EU is in violation of the SPS Agreement. Moreover, the EU has ignored the available scientific information regarding CDC Triffid flax. The EU did not consider the possibility of following the guidelines given by Codex Alimentarius Commission in the case of CDC Triffid flax. There are non-scientific reasons behind the EUs zero tolerance on CDC Triffid flax and they overweigh the available scientific information. The EU position would be unlikely to be supported if a complaint was brought to the World Trade Organisation Disputes Panel. A partial equilibrium model was used to provide a theoretical background to examine the changes in the flaxseed industry and the linseed oil industry due to the CDC Triffid event. A model of the supply chain of Canadian flaxseed was developed to illustrate the operationalisation of the Protocol developed by the EU and Canada to address the zero tolerance policy. Empirical estimation suggests that the operationalisation of the Protocol incurred additional cost of $7.5 million to the flax seed industry of Canada in 2009/ 2010. Out of that, cost of testing was approximately $1.2 million and cost of segregation was $4.2 million. Estimation of changes in revenue suggests that there was a loss of revenue in flaxseed trade between the EU and Canada in 2009/2010. Imports of Canadian flax by China provided an alternative market, at a considerably lower price than typically realised from the EU market. Interestingly, the EUs zero tolerance policy on CDC Triffid flax has resulted in a larger additional cost on the EU than Canada.

Flax is one of the major cash crops in Canada. Approximately seventy percent of Canadian flaxseed was exported to European Union (EU) annually until 2009. In 2009, the EU imposed an import ban on Canadian flaxseed due to the adventitious presence of a GM flax variety - CDC Triffid was identified in Canadian flaxseed exported to the EU. The EUs decision to apply zero tolerance on CDC Triffid flax has been based on its interpretation of the precautionary principle. According to the World Trade Organisations Agreement on the Application of Sanitary and Phytosanitary Measures (SPS), however, precautionary measures are subject to a scientific risk assessment. As the EU did not base its zero tolerance for CDC Triffid flax on any scientific risk assessment, the EU is in violation of the SPS Agreement. Moreover, the EU has ignored the available scientific information regarding CDC Triffid flax. The EU did not consider the possibility of following the guidelines given by Codex Alimentarius Commission in the case of CDC Triffid flax. There are non-scientific reasons behind the EUs zero tolerance on CDC Triffid flax and they overweigh the available scientific information. The EU position would be unlikely to be supported if a complaint was brought to the World Trade Organisation Disputes Panel. A partial equilibrium model was used to provide a theoretical background to examine the changes in the flaxseed industry and the linseed oil industry due to the CDC Triffid event. A model of the supply chain of Canadian flaxseed was developed to illustrate the operationalisation of the Protocol developed by the EU and Canada to address the zero tolerance policy. Empirical estimation suggests that the operationalisation of the Protocol incurred additional cost of $7.5 million to the flax seed industry of Canada in 2009/ 2010. Out of that, cost of testing was approximately $1.2 million and cost of segregation was $4.2 million. Estimation of changes in revenue suggests that there was a loss of revenue in flaxseed trade between the EU and Canada in 2009/2010. Imports of Canadian flax by China provided an alternative market, at a considerably lower price than typically realised from the EU market. Interestingly, the EUs zero tolerance policy on CDC Triffid flax has resulted in a larger additional cost on the EU than Canada.