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Resumo Dyckia distachya é uma bromélia endêmica terrestre nativa do Brasil e ameaçada de extinção uma vez que seu habitat natural, a Mata Atlântica, encontra-se extremamente vulnerável. Por ser uma bromélia terrestre, suas raízes são funcionais e absorvem nutrientes do solo. Este trabalho estudou duas formas de indução ao enraizamento, in vitro e ex vitro, utilizando como explante inicial, brotações micropropagadas. Os resultados obtidos mostraram que o enraizamento das brotações de D. distachya deve ser realizado in vitro com a utilização de AIB nas concentrações de 10-15 μM por 10-15 dias e posterior expressão em meio isento de AIB, antecedendo a transferência para as condições ex vitro.

Eugenia involucrata DC. (Myrtaceae), es un árbol nativo de Argentina, Brasil, Paraguay y Uruguay, con importante potencial económico. Su propagación y conservación están limitados por la característica recalcitrante de sus semillas, siendo el cultivo de tejidos un método eficiente para estos fines. En este trabajo se evaluó el efecto del tipo de explante y distintas combinaciones de reguladores de crecimiento vegetal, BAP, TDZ y ANA en la formación de brotes adventicios, enraizamiento y formación de plántulas de E. involucrata. Se desarrollaron tres experimentos: inducción, diferenciación de brotes y aclimatización en invernáculo de las plántulas obtenidas. Las hormonas BAP y TDZ indujeron callos y brotes a partir de segmentos nodales, radiculares y foliares. El máximo porcentaje de formación de brotes por organogénesis directa se obtuvo en segmentos nodales cultivados en medio MS ½ con 2,25 mg/L de BAP y la máxima frecuencia de formación de callos se obtuvo en explantos nodales o radiculares cultivados en medio MS ½ con TDZ, solo o combinado con ANA. La máxima formación de brotes adventicios, por organogénesis indirecta, se obtuvo en segmentos nodales, inducidos en medio MS ½ con 1,1 mg/l de TDZ+0,93 mg/l de ANA y subcultivados en medio MS ½ con 0,5 mg/l de TDZ +1 g/l de carbón activado. Los ápices de los brotes obtenidos a partir de organogénesis directa e indirecta enraizaron in vitro en medio MS ½ libre de RCV formando plántulas que luego se aclimataron en invernáculo, demostrando un alto porcentaje de supervivencia a los seis meses de trasplante.

Pineapple (Ananas comosus) is a horticultural species of the Bromeliaceae family of high socioeconomic interest, widely cultivated around the world. The multiplication of pineapple seedlings in the field can be time-consuming, requiring a significant labor investment. The objective of this study was to evaluate the performance of continuous and temporary immersion systems in the micropropagation scale-up of the species. Shoots were obtained from explants subcultured in flasks with gelled culture medium and without gas exchange. The shoots were transferred to liquid MS medium supplemented with 2mM NAA, and 4mM BAP, and cultivated in four different devices: sealed flasks, flasks with semipermeable gas membranes, RITA®, and twin-flasks. After 45 days of cultivation, plant growth, fresh mass increment, the stomatal density of the abaxial surface of the leaves, the maximum quantum yield of the photosystem II, and contents of chlorophyll and carotenoids were analyzed. Significant differences were observed in plant growth, stomatal density, and contents of chlorophyll and carotenoids. The twin-flasks and RITA® devices revealed better results in morphological parameters, such as plant growth and stomatal density, while the treatments in sealed flasks and with membrane stood out in the chlorophyll content.

Somatic embryogenesis (SE) in conifers is usually characterized as a multi-step process starting with the development of proembryogenic cell masses and followed by histodifferentiation, somatic embryo development, maturation, desiccation, and plant regeneration. Our current understanding of conifers’ SE is mainly derived from studies using Pinaceae species as a model. However, the evolutionary relationships between conifers are not clear. Some hypotheses consider conifers as a paraphyletic group and Gnetales as a closely related clade. In this review, we used an integrated approach in order to cover the advances in knowledge on SE in conifers and Gnetales, discussing the state-of-the-art and shedding light on similarities and current bottlenecks. With this approach, we expect to be able to better understand the integration of these clades within current studies on SE. Finally, the points discussed raise an intriguing question: are non-Pinaceae conifers less prone to expressing embryogenic competence and generating somatic embryos as compared to Pinaceae species? The development of fundamental studies focused on this morphogenetic route in the coming years could be the key to finding a higher number of points in common between these species, allowing the success of the SE of one species to positively affect the success of another.

ABSTRACT Although karyotype features are useful data for evolutionary studies, cytogenetic data in Bambusoideae are mainly based only on chromosome counts. The first comparative cytogenetic analysis of three Neotropical woody bamboo species - Guadua chacoensis, G. angustifolia and Chusquea tenella - was undertaken based on new and reviewed chromosome counts, CMA/DAPI double staining, fluorescent in situ hybridization (FISH) with 35S and 5S rDNA probes, and genome size estimation. The two species of Guadua were found to have 2n = 46 chromosomes, while the first record for C. tenella was 2n = 44. Only one pair of CMA+/DAPI- was detected on the terminal region of metacentric chromosomes in all three species. Likewise, one pair of 5S and 35S rDNA sites was detected in all three species, with the 35S rDNA sites always collocated with the CMA+ bands. Genome sizes ranged from 2C ≈ 3.99 pg for the species of Guadua, to 2C = 4.77 pg for C. tenella. Considering the Miocene origin of Neotropical woody bamboos, the observed karyotype stability suggests that the analyzed species are diploidized paleopolyploids. The results reveal the conservative cytomolecular organization of Neotropical woody bamboo karyotypes, which helps to improve our understanding of the evolution of this group.

Eugenia uniflora is an American tree species with ecological, agronomical and medicinal importance. However, just few studies have focused on the in vitro propagation and production of secondary metabolites. This study investigated the explant sources and culture conditions for the in vitro callogenesis in E. uniflora towards induction of somatic embryogenesis and production of antioxidant compounds. Nodal segments, leaf sections and root segments from in vitro germinated seeds were used as explants and eight different combinations of auxins (2,4-D or NAA) and cytokinins (BAP or TDZ) were tested for the callus induction. The best callogenic response was observed in nodal segments, followed by leaf sections. Root segments presented comparatively poorer callogenic performance. Calli from nodal segments cultivated in MS medium with NAA (10 μM) + TDZ (5 μM) originated putative pro-embryogenic structures, while the culture in liquid medium using MS supplemented with NAA (10 μM) + TDZ (5 μM) revealed significantly higher content of phenols and flavonoids, as well as higher reducing capacity than the tested treatments and the control (fresh leaves). In summary, the calli obtained from nodal segments revealed competence for somatic embryogenesis induction and development as well as the production of secondary metabolites with pharmaceutical potential.

Este trabalho teve por objetivo apresentar indicadores técnicos e econômicos para o manejo sustentado do palmiteiro (Euterpe edulis M.) no Estado de Santa Catarina, a partir de uma unidade demonstrativa de manejo implantada na Floresta Nacional de Ibirama, SC – Flona de Ibirama, SC.

This study aimed to establish a cryopreservation protocol for G. chacoensis embryogenic cultures (ECs) and to investigate the role of antioxidant enzymes activities during cryopreservation. The growth dynamics of cell suspensions were also investigated, followed by a phytotoxicity test to assess the ECs’ ability to tolerate the use of cryoprotective solutions for different incubation times (0, 30, 60, 120, and 240 min). We evaluated the EC redox state in three steps of cryopreservation: after incubation in cryoprotection solution, after thawing, and 60 days after regrowth. Our results showed that the ECs support the use of cryoprotective solution until 120 min, showing phytotoxic effects with 240 min of incubation. This study reports a 100% survival of the cultures and a 10% increase ratio in fresh material for both incubation times tested (60 and 120 min). Increased malonaldehyde content was identified after incubation in the cryoprotective solution. An increase in the activities of catalase and ascorbate peroxidase was also identified in the subsequent steps, suggesting that the activation of antioxidant enzymes is essential for maintaining cell homeostasis during cryopreservation.

Changes in the chemical environment at the maturation stage in Pinus spp. somatic embryogenesis will be a determinant factor in the conversion of somatic embryos to plantlets. Furthermore, the study of biochemical and morphological aspects of the somatic embryos could enable the improvement of somatic embryogenesis in Pinus spp. In the present work, the influence of different amino acid combinations, carbohydrate sources, and concentrations at the maturation stage of Pinus radiata D. Don and Pinus halepensis Mill. was analyzed. In P. radiata, the maturation medium supplemented with 175 mM of sucrose and an increase in the amino acid mixture (1,100 mgL–1 of L-glutamine, 1,050 mgL–1 of L-asparagine, 350 mgL–1 of L-arginine, and 35 mgL–1 of L-proline) promoted bigger embryos, with a larger stem diameter and an increase in the number of roots in the germinated somatic embryos, improving the acclimatization success of this species. In P. halepensis, the maturation medium supplemented with 175 mM of maltose improved the germination of somatic embryos. The increase in the amount of amino acids in the maturation medium increased the levels of putrescine in the germinated somatic embryos of P. halepensis. We detected significant differences in the amounts of polyamines between somatic plantlets of P. radiata and P. halepensis; putrescine was less abundant in both species. For the first time, in P. radiata and P. halepensis somatic embryogenesis, we detected the presence of cadaverine, and its concentration changed according to the species.

O morango cultivado (Fragaria x ananassa Duch. (Rosaceae) é um híbrido originado pelo cruzamento das espécies americanas Fragaria chiloensis e Fragaria virginiana e pertence à família Rosaceae. O morangueiro possui reprodução vegetativa e, por isso, é comum o acúmulo de viroses e outras doenças de difícil controle. Uma das quatro viroses mais importantes na cultura do morango é causada pelo vírus da faixa das nervuras (Strawberry vein banding virus – SVBV), que é transmitido no campo por afídeos, de maneira semipersistente. Para melhorar o conhecimento genômico, são recomendadas técnicas moleculares e a classificação de isolados de SVBV. Um dos determinantes primários da transmissibilidade e especificidade por afídeos é a proteína do capsídeo, que possui importância crítica para o estabelecimento da infecção. Neste trabalho, foi sequenciada parcialmente a proteína do capsídeo do gene do SVBV de um isolado alemão, inoculado em morangueiros e mantido em casa de vegetação no Brasil, por mais de dez anos. Foi realizada a análise filogenética comparando as sequências contidas no GenBank com o objetivo de elucidar as relações evolutivas nesta espécie. As análises filogenéticas mostraram que a sequência do isolado está mais próxima dos isolados dos EUA e Egito. Estes resultados contribuem para a melhor elucidação dos mecanismos de evolução do vírus e do patossistema em questão.

Profiling technologies, such as proteomics, allow the simultaneous measurement and comparison of thousands of plant components without prior knowledge of their identity. The combination of these non-targeted methods facilitates a more comprehensive approach than targeted methods and thus provides additional opportunities to identify genotypic changes resulting from genetic modification, including new allergens or toxins. The purpose of this study was to investigate unintended changes in GM Bt maize grown in South Africa. In the present study, we used bi-dimensional gel electrophoresis based on fluorescence staining, coupled with mass spectrometry in order to compare the proteome of the field-grown transgenic hybrid (MON810) and its near-isogenic counterpart. Proteomic data showed that energy metabolism and redox homeostasis were unequally modulated in GM Bt and non-GM maize variety samples. In addition, a potential allergenic protein—pathogenesis related protein −1 has been identified in our sample set. Our data shows that the GM variety is not substantially equivalent to its non-transgenic near-isogenic variety and further studies should be conducted in order to address the biological relevance and the potential risks of such changes. These finding highlight the suitability of unbiased profiling approaches to complement current GMO risk assessment practices worldwide.

Estudou-se o estado fisiológico das sementes de várias espécies mantidas nas coleções de germoplasma da Epagri por meio de testes de viabilidade, germinação e índice de vigor. Dez acessos foram amostrados para cada uma das espécies: Phaseolus vulgaris, Allium cepa, Oriza sativa, Paspalum dilatatum, Bromus auleticus, Trifolium repens e Adesmia tristis. A análise de dados indicou que os métodos de avaliação se prestam ao monitoramento dos acessos, cujas sementes, nestas condições, apresentaram longevidade estimada entre 18 e 32 anos e degradação fisiológica dependente da espécie e do ambiente de armazenagem.

Dando continuidade ao seu programa de melhoramento da goiabeira serrana (Acca selowiana), uma das fruteiras nativas mais promissoras do Sul do Brasil em termos comerciais, a Epagri está lançando duas novas cultivares, SCS 414-Mattos e SCS 415-Nonante, que têm por objetivo estender e melhorar o fluxo de oferta de goiabas serranas ao mercado. O pico de colheita da cultivar Mattos encaixa-se entre o da ‘Alcântara’ e da ‘Helena’, lançadas em 2007, enquanto ‘Nonante’ começa a amadurecer cerca de 20 dias após o início da colheita da ‘Helena’, estendendo o período de colheita da goiaba serrana até a segunda semana de maio. A cultivar Mattos resulta da seleção e propagação vegetativa de um dos 200 acessos silvestres do banco de germoplasma e destacou-se pelo tamanho, aparência e qualidade do fruto, enquanto ‘Nonante’ resulta de um cruzamento entre dois acessos coletados em quintais de Santa Catarina, e se destacou pela constância de produção, rusticidade e sabor de seus frutos, além de ser autocompatível.

Abstract Butia eriospatha is an endemic palm species from the Atlantic Rainforest in Brazil, a biodiversity hotspot. This species is currently listed in the IUCN red list as vulnerable and lacks specific plastid markers for population genetics studies. In addition, the evolutionary relationship within the genus Butia is not yet well resolved. Here, we sequenced and characterized the complete plastid genome (plastome) sequence of B. eriospatha. The complete plastome sequence is 154,048 bp in length, with the typical quadripartite structure. This plastome length and genes content is consistent with other six species from tribe Cocoseae. However, the Inverted Repeat (IR) borders show some variation among the analyzed species from this tribe. Species from the Bactridinae (Astrocaryum and Acrocomia) and Elaeidinae (Elaeis) subtribes present the rps19 gene completely duplicated in the IR region. In contrast, all plastomes sequenced from the subtribe Attaleinae (Butia, Cocos, Syagrus) present one complete CDS of rps19 and one partial copy of rps19. The difference in the IR/LSC junctions between Attaleinae and the sister clades Bactridinae + Elaeidinae might be considered an evolutionary signal and the plastome sequence of B. eriopatha may be used in future studies of population genetics and phylogeny.

Tissue culture techniques have been employed for bromeliad mass propagation by means of the morphogenetic route of nodular cluster cultures (NCs). This study aimed to assess protein, carbohydrate and global DNA methylation (GDM) level dynamics during NCs induction from Vriesea reitzii seeds. Seeds were inoculated into Murashige and Skoog (MS) liquid medium supplemented with 4 µM α-naphthaleneacetic acid (NAA) to induce NCs and in culture medium without plant growth regulators to form normal seedlings. Samples collected at 0, 3, 7, 10, 14, and 21 days of culture were analyzed. All parameters assessed showed the same variation pattern. However, seeds inducing NCs showed significantly lower starch (6.0 mg g-1 FM), carbohydrate (10.7 mg g-1 FM) and GDM (11.0%) levels than seeds forming normal seedlings after 21 days in culture. On the other hand, the protein content (9.1 mg g-1 FM) was significantly higher during induction. NCs induction process through seeds is the result of gene reprogramming in the explant, which leads to morphological, biochemical and metabolic alterations. This involves dedifferentiation, high cell proliferation, high energy demand and protein synthesis, which is related to elevated metabolic activity.

While some genetically modified (GM) plants have been targeted to confer tolerance to abiotic stressors, transgenes are impacted by abiotic stressors, causing adverse effects on plant physiology and yield. However, routine safety analyses do not assess the response of GM plants under different environmental stress conditions. In the context of climate change, the combination of abiotic stressors is a reality in agroecosystems. Therefore, the aim of this study was to analyze the metabolic cost by assessing the proteomic profiles of GM soybean varieties under glyphosate spraying and water deficit conditions compared to their non-transgenic conventional counterparts. We found evidence of cumulative adverse effects that resulted in the reduction of enzymes involved in carbohydrate metabolism, along with the expression of amino acids and nitrogen metabolic enzymes. Ribosomal metabolism was significantly enriched, particularly the protein families associated with ribosomal complexes L5 and L18. The interaction network map showed that the affected module representing the ribosome pathway interacts strongly with other important proteins, such as the chloro-plastic gamma ATP synthase subunit. Combined, these findings provide clear evidence for increasing the metabolic costs of GM soybean plants in response to the accumulation of stress factors. First, alterations in the ribosome pathway indicate that the GM plant itself carries a metabolic burden associated with the biosynthesis of proteins as effects of genetic transformation. GM plants also showed an imbalance in energy demand and production under controlled conditions, which was increased under drought conditions. Identifying the consequences of altered metabolism related to the interaction between plant transgene stress responses allows us to understand the possible effects on the ecology and evolution of plants in the medium and long term and the potential interactions with other organisms when these organisms are released in the environment.

The contamination by microorganisms and oxidation of explants in the in vitro establishment of bamboo are recurrent problems for its micropropagation. In the present study, effects of the biocide Plant Preservative Mixture (PPM™), the antibiotic kasugamycin and the consistency of the culture medium were evaluated in the in vitro establishment of Bambusa vulgaris,Phyllostachys bambusoides and Dendrocalamus asper. The presence of PPM™ in the culture medium had a significant effect using 2 mL L-1 or 4 mL L-1 concentrations, as well as in the liquid culture medium, increasing the plants established in the autumn. Kasugamycin promoted variable responses for all the three species, depending on the season. There was interaction among the factors, demonstrating that higher rates of viable plants can be obtained by combining different strategies to reduce the oxidation and contamination. For the in vitro establishment of B. vulgaris,P. bambusoides and D. asper, it is recommended to add 2 mL L-1 or 4 mL L-1 of PPM™ to the liquid culture medium.

The bamboo productive chain is still incipient in Brazil, and the low supply of plantlets due to low-efficient conventional propagation methods presents a significant bottleneck to its development. This study aimed to establish a micropropagation protocol for Guadua chacoensis. Explants from donor plants cultivated under controlled environment showed less contamination, if compared to explants from plants grown in the field. The contamination rate was even lower when 2 mL L-1 of Plant Preservative Mixture (PPM™) were added to the culture medium, leading to a higher establishment rate. The obtained cultures were then multiplied using either in vitro-derived nodal segments or clump division in the presence of increasing contents (0 mM, 10 mM, 20 mM, 30 mM or 40 mM) of 6-Benzylaminopurine (BAP). The number of shoots increased with increasing BAP concentrations, but this also resulted in a reduced rooting rate and root length. Plants acclimatized under 0 %, 35 % or 65 % of shading showed a dynamic maximum quantum yield of photosystem II (Fv/Fm), which initially decreased within the first seven days after the transfer to ex vitro conditions, but then increased until reaching stable values of 0.775 after 17 days. Additionally, the shading improved the plant survival rates, if compared to those under non-shaded conditions, which presented photoinhibition and photodamage symptoms.

In this study, an efficient method for regenerating plants from nodal cultures of seedlings was developed for Aspidosperma polyneuron. Mature seeds were surface-sterilized and embryos were germinated in Woody Plant Medium (WPM). Epicotyl and hypocotyl nodal segments, excised from 3-week-old in vitro -grown seedlings, were cultured in WPM medium supplemented with 6-benzyladenine (BA) (2.5, 5.0, and 10 μM), alone or combined with indole-3-butyric acid (IBA) or α-naphthaleneacetic acid (NAA) (0.5 μM) for culture initiation and three subcultures. For root induction, IBA (2.5, 5.0, and 10 mM) pulse treatments (15 minutes) were initially applied, followed by transfer to growth regulator-free WPM for five weeks. Regenerated plants were first transplanted to trays containing soil and Plantmax ® substrate (3:1) and later to polyethylene bags for acclimatization in a greenhouse. Hypocotyl explants exhibited superior shoot regeneration rate and rooting percentages compared with those of epicotyl explants. The highest numbers of shoots per explant (7 −8) were obtained at the third subculture when using the culture medium supplemented with 10 μM BA. When combined with BA, the addition of 0.5 μM IBA or NAA did not affect the regeneration of shoots. An IBA pulse treatment of 5 − 10 mM for 15 minutes induced 60% of rooting. The regenerated plants were acclimatized and successfully established in a greenhouse, with a 90% survival rate observed after three months. Based on the results of this study, the micropropagation of Aspidosperma polyneuron from in vitro seedlings is feasible and could be a useful tool for the conservation and propagation of this important endangered species.

Climate change will inevitably lead to environmental variations, thus plant drought tolerance will be a determinant factor in the success of plantations and natural forestry recovery. Some metabolites, such as soluble carbohydrates and amino acids, have been described as being the key to both embryogenesis efficiency and abiotic stress response, contributing to phenotypic plasticity and the adaptive capacity of plants. For this reason, our main objectives were to evaluate if the temperature during embryonal mass initiation in radiata pine was critical to the success of somatic embryogenesis, to alter the morphological and ultrastructural organization of embryonal masses at cellular level and to modify the carbohydrate, protein, or amino acid contents. The first SE initiation experiments were carried out at moderate and high temperatures for periods of different durations prior to transfer to the control temperature of 23°C. Cultures initiated at moderate temperatures (30°C, 4 weeks and 40°C, 4 days) showed significantly lower initiation and proliferation rates than those at the control temperature or pulse treatment at high temperatures (50°C, 5 min). No significant differences were observed either for the percentage of embryogenic cell lines that produced somatic embryos, or for the number of somatic embryos per gram of embryonal mass. Based on the results from the first experiments, initiation was carried out at 40°C 4 h; 50°C, 30 min; and a pulse treatment of 60°C, 5 min. No significant differences were found for the initiation or number of established lines or for the maturation of somatic embryos. However, large morphological differences were observed in the mature somatic embryos. At the same time, changes observed at cellular level suggested that strong heat shock treatments may trigger the programmed cell death of embryogenic cells, leading to an early loss of embryogenic potential, and the formation of supernumerary suspensor cells. Finally, among all the differences observed in the metabolic profile, it is worth highlighting the accumulation of tyrosine and isoleucine, both amino acids involved in the synthesis of abiotic stress response-related secondary metabolites.

The present study intended to investigate the effects of different glutathione (GSH) levels (0, 0.1, 0.5 and 1 mM) on the somatic embryogenesis (SE) induction of Acca sellowiana. Besides, we evaluated the effect of different carbon sources (sucrose and maltose) on the somatic embryos conversion. GSH-supplemented treatments resulted in improved SE induction rates (~70%) as compared to the control GSH-free (~35%) after 50 days of culture. The total number of somatic embryos obtained did not differ between treatments, but significant differences were observed for the embryonic stages after 80 days of culture. After 80 days of culture, 0.5 and 1 mM GSH-supplemented treatments showed the largest amount of torpedo-staged somatic embryos. In contrast, treatments supplemented with 0 and 0.1 mM GSH showed equal amounts of somatic embryos at all embryonic stages. These results indicate that GSH accelerates the SE induction process and increases the synchrony of the somatic embryo formation of A. sellowiana. The use of maltose for the somatic embryos conversion, as compared to sucrose, did not influence the conversion rate of normal chlorophyllous somatic embryos, but increased the formation of normal achlorophyllous somatic plantlets. This finding can be attributed to the rapid hydrolysis of sucrose, contributing to an enhanced chlorophyll synthesis.

A agrobiodiversidade é a parcela da biodiversidade constituída por um conjunto de organismos e ecossistemas que apresentam fortes relações com os seres humanos, podendo ser domesticados, semidomesticados, cultivados ou manejados pelo homem. Essa diversidade, conservada há séculos pelo uso (conservação in situ ou conservação on farm) pelos pequenos agricultores em todas as regiões do planeta, encontra-se ameaçada. De um lado, a agricultura industrial ou química através de suas práticas, ricas em insumos químicos, provoca muitas externalidades negativas, até recentemente pouco consideradas. De outro lado, as mudanças climáticas, igualmente, produzem efeitos adversos à diversidade genética. No entanto, as preocupações com a produção de alimentos de alta qualidade biológica em sistemas agrícolas sustentáveis remontam ainda o início do século XX. Dentre os vários sistemas agrícolas sustentáveis ou alternativos, a agroecologia apresenta vários atributos favoráveis: não faz uso de insumos químicos, é sustentável em todas as suas dimensões, faz uso de grande diversidade genética no cultivo, é socioeconômico-associada, estimula as relações de vizinhança e produz alimentos com alta qualidade biológica e nutricional. No âmbito da agroecologia e não da agricultura industrial ou química, há inúmeras oportunidades para a C&T desenvolver pesquisas participativas, contextualizadas, que podem empoderar tanto a agricultura familiar quanto as comunidades tradicionais no aperfeiçoamento dos processos e princípios agroecológicos utilizados. Assim, muitas das externalidades negativas atualmente inadmissíveis poderiam ser evitadas.

Micropropagation based techniques enable the mass production of pest- and disease-free stock plants. Aiming at optimizing pineapple [Ananas comosus (L.) Merril] micropropagation, a complete randomized experiment with three factors was established: immersion system (2 L twin flasks temporary immersion and permanent immersion); culture medium (supplemented or not with GA3 ); and shoot height class. The effects on regeneration rate (number of new shoots per 1 g fresh mass (FM) of inoculated microshoots), FM per shoot, and shoot survival were evaluated. Significant interactions (p

Tissue culture techniques have been employed for orchid mass propagation by means of the morphogenetic route of protocorm like-bodies (PLBs). This study aimed to analyze and compare Indole-acetic acid (IAA) and sugar endogenous levels in protocorm-like bodies (PLBs) induction and development in Cattleya tigrina, in order to better understand this process and to optimize micropropagation procedure protocols. Leaves grown on MS (Murashige and Skoog) culture medium, suplemented with 9 μM TDZ for PLBs induction and development were collected after 0, 2, 7, 14, 30, 60 and 100 days of cultivation, for further analysis. Increase of IAA and reduction of sugar levels are strongly related to morphogenetic response, that is, PLBs formation over the preexisting ones and leaf primordia formation. Sucrose, fructose and glucose presence in this study is related to cell signaling. Thus, hormonal signals and carbohydrates alter metabolism, triggering PLBs initiation and development in C. tigrina.

Cattleya tigrina is endemic to the Atlantic forest biome and classified as vulnerable in the Red Book of Brazilian Flora. In vitro techniques comprise valuable tools for the conservation of endangered plant species. The aim of the present study was to evaluate the morphological features, global DNA methylation levels and free polyamines during protocorm-like bodies (PLBs) induction of C. tigrina. Along with that, an efficient protocol for in vitro propagation of this species is proposed. The first evidences of PLBs induction in C. tigrina occurred at seven days in culture, starting from the basal portion of the leaf abaxial surface. A hypomethylation marked the beginning of cell differentiation, followed by an increased global DNA methylation at 35 days in culture, coinciding with a subtle change in the structures morphogenetic development. During PLBs induction, putrescine exhibited higher levels as compared to spermidine and spermine, and apparently presents a major role during the PLBs induction in C. tigrina. Due to the apparent secondary PLBs formation, this protocol can represent a highly efficient method for in vitro propagation of this species.

The asymbiotic technique of orchid seeds germination is an important method of mass production of seedlings. Studies on the best culture conditions for each species are important to obtain seedlings in less time and at lower costs. Current analysis evaluates different consistencies of culture medium, saline formulations and culture conditions on the germination rate and further development of protocorms of Epidendrum fulgens. After 45 days in culture the protocorms were classified into three categories of development. The liquid saline formulation of Murashige and Skoog (1962) (MS) provided the highest germination rate (83.5%), and the Knudson formulation (1946) the lowest (10.9%). The different consistencies or conditions or culture conditions did not affect the germination rate percentage, except the Knudson medium, which resulted in the highest rate in response to the gelled consistency. Protocorms cultured in liquid MS medium with or without agitation showed the fastest development.

The induction of nodular culture (NC) and the subsequent development of microshoots of V. reitzii are considered an in vitro propagation model-system with high regenerative performance. Current research analyzed the determinant factors of the in vitro morphogenesis control of bromeliads. Seeds excised from mature capsules were grown on medium MS basic (MSB), liquid or gelled, supplemented or not with α-naphthaleneacetic acid (NAA), 6-benzilaminopurine (BAP) or thidiazuron (TDZ). The regeneration and elongation of microshoots were evaluated from NC sub-cultivated on MSB medium on liquid culture medium supplemented with different concentrations of indolyl-3-acetic acid (IAA) and gibberellic acid (GA3). Plant growth regulators (PGR) supplemented into the medium MSB inhibited the germination of the seeds and induced NC in the second week of growth. The induced NC on MSB medium with NAA (4 µM) and sub-cultivated on MSB medium with NAA (2 µM) plus N6(2-isopentenyl) adenine (2-iP) (2 µM) showed granular texture and high rate of proliferation. NC sub-culture in MSB medium with IAA (4 µM) provided a higher average number of microshoots (1,478 shoots g-1 of NC). Shoots over 3.0 cm resulted in more than 95% ex vitro survival.

Angiosperm and gymnosperm plants evolved from a common ancestor about 300 million years ago. Apart from morphological and structural differences in embryogenesis and seed origin, a set of embryogenesis-regulating genes and the molecular mechanisms involved in embryo development seem to have been conserved alike in both taxa. Few studies have covered molecular aspects of embryogenesis in the Brazilian pine, the only economically important native conifer in Brazil. Thus eight embryogenesis-regulating genes, viz.,ARGONAUTE 1, CUP-SHAPED COTYLEDON 1, WUSCHEL-related WOX, S-LOCUS LECTIN PROTEIN KINASE, SCARECROW-like, VICILIN 7S, LEAFY COTYLEDON 1, and REVERSIBLE GLYCOSYLATED POLYPEPTIDE 1, were analyzed through semiquantitative RT-PCR during embryo development and germination. All the eight were found to be differentially expressed in the various developmental stages of zygotic embryos, seeds and seedling tissues. To our knowledge, this is the first report on embryogenesis-regulating gene expression in members of the Araucariaceae family, as well as in plants with recalcitrant seeds.

Kiwi is a dioecious plant species, requiring cross pollination for fruit production. The objective of this study was toevaluate the in vitro viability and shelf life of pollen grains of two kiwi varieties. Flowers of the cultivars Matua and Tomuri werecollected and the pollen germinated in vitro, in culture medium containing agar (1 %), sucrose (0, 5, 10, 20 and 40 %) and boric acid(0 and 50 mg L-1H3BO3). Pollen grains were stored in a BOD incubator (25.0 °C), refrigerator (4.0 °C), freezer (-18.0 °C) and in liquid N2 (-196.0 °C), and evaluated after 0, 40, 120, 240 and 365 days. The culture medium enriched with 12 % sucrose and 50 mgL-1 H3BO3 was the most suitable. Pollen grains can be stored for a short period in the refrigerator or freezer, and cryopreserved for at least one year.

A variedade Goethe é símbolo da vitivinicultura da região de Urussanga, sul do Estado de Santa Catarina, a qual, atualmente, busca a Indicação Geográfica da Uva e do Vinho Goethe. Para isto, um dos requisitos necessários é a identificação precisa do material genético. Os marcadores microssatélites constituem a ferramenta molecular mais utilizada para a identificação varietal de videira em todo o mundo e têm a capacidade de produzir um perfil genético único para cada material vitícola. O objetivo deste trabalho foi caracterizar duas seleções de uva 'Goethe', presentes no município de Urussanga, por meio de marcadores moleculares microssatélites, visando a atender aos requisitos de denominação de origem e indicação de procedência controlada. A extração do DNA genômico foi realizada a partir de folhas jovens e ramos de nove acessos de cada seleção de 'Goethe Classica' e 'Goethe Primo' provenientes de uma coleção pública e de oito coleções privadas da região de Urussanga. Dez loci microssatélites VVS2, VVMD5, VVMD7, VVMD27, VrZAG62, VrZAG79, VVMD25, VVMD28, VVMD31 e VVMD32 foram genotipados através de eletroforese capilar. As análises realizadas mostraram que as duas variantes da uva 'Goethe' apresentaram um perfil molecular idêntico e único, isto é, representam a mesma variedade e sem nenhuma correspondência com variedades descritas anteriormente na literatura e nos bancos de dados consultados. As diferenças fenotípicas observadas provavelmente são devidas a mutações somáticas em regiões funcionais do genoma, fenômeno que dá origem aos clones em videira.'Goethe' grape is a symbol of the viticulture of Urussanga region, South of Santa Catarina State, which is currently claiming the geographical indication of the grape and wine Goethe. Microsatellite markers are the biotechnological tool most used for molecular identification of grapevine varieties worldwide. These markers have the potential of generate unique DNA profiles for all the diverse grape genotypes. The aim of this work was to characterize two selections of the grapevine 'Goethe' from Urussanga, using microsatellite markers. Genomic DNA was extracted from young leaves and branches of nine accessions of each selection "Goethe Clássica and 'Goethe Primo' maintained at one public and eight private collections of Urussanga. Ten microsatellite loci VVS2, VVMD5, VVMD7, VVMD27, VrZAG62, VrZAG79, VVMD25, VVMD28, VVMD31 and VVMD32 were genotyped by capillary electrophoresis. Results showed that the two variants of 'Goethe' have an identical molecular profile which doesn't match any other DNA profile of grapevines previously described in literature or available in databases. The phenotypic differences observed are probably due to somatic mutations in functional regions of the genome, a phenomenon which leads to the creation of clones in grape.

O presente trabalho objetivou estabelecer um protocolo para obtenção de culturas assépticas de Aspidosperma polyneuron visando à regeneração in vitro de mudas dessa espécie. Brotações apicais de mudas de dois anos de idade foram coletadas em casa de vegetação e desinfestadas com hipoclorito de sódio (NaOCl) ou cloreto de mercúrio (HgCl2) visando ao estabelecimento de culturas assépticas. Os tratamentos testados com NaOCl foram: solução 0,125 ou 0,25%, durante cinco ou dez minutos e com HgCl2: 0,025; 0,05 ou 0,1%, durante cinco ou dez minutos e em todas as estações do ano. As avaliações das percentagens de necrose, contaminação fúngica, contaminação bacteriana e de sobrevivência foram feitas após três semanas. Os resultados revelaram que o tratamento com solução de NaOCl a 0,25%, durante 10 minutos, resultou em 70% de sobrevivência para brotações apicais, independente da época do ano (verão e outono). O HgCl2 foi mais eficiente que o NaOCl na desinfestação de brotações apicais de peroba-rosa, sendo recomendado o tratamento de 0,05% de HgCl2, durante 10 minutos (84,10% de sobrevivência). Em todas as estações do ano, foram estabelecidas culturas assépticas, contudo, os melhores resultados foram obtidos na primavera e verão.

Embryogenic cultures of Araucaria angustifolia were established in a BM liquid medium supplemented with 2 µM 2,4-dichlorophenoxyacetic acid, 1 µM 6-benzylaminopurine and 1 µM kinetin (BM2) and in a BM medium free of growth regulators (BM0). During 42 days in culture, the cell growth pattern of both cultures was similar. The pH of the culture medium of both BM0 and BM2 underwent progressive reduction during culture time. For both the embryogenic cultures a preferential uptake of glucose in the late stages of cell growth kinetics was observed. The extracellular protein content was similar for both the embryogenic cultures. Acetocarmine and Evan's blue double stain showed major differences for early somatic embryo organisation, in which only the embryogenic culture grown in a liquid culture medium free of plant growth regulators showed the presence of bipolar somatic pro-embryos.Culturas embriogênicas de Araucaria angustifolia foram estabelecidas em meio de cultura líquido BM suplementado com 2 µM Ácido 2,4 Diclorofenoxiacético, 1 µM 6-Benzilaminopurina e 1 µM Cinetina (BM2) e em meio BM isento de reguladores de crescimento (BM0). Durante 42 dias de cultivo, o padrão de crescimento celular em ambas as culturas foi similar. O pH do meio de cultura BM0 e BM2 sofreu uma progressiva redução durante o período de cultivo. Em ambas as culturas embriogênicas foram observadas um consumo preferencial de glicose no período final da curva de crescimento celular. O nível de proteínas extracelulares foi similar para ambas as culturas embriogênicas. A dupla coloração com carmin acético e azul de Evans revelou diferenças na organização das linhagens celulares embriogênicas, sendo que a presença de proembriões somáticos bipolares foi apenas evidenciada nas culturas embriogênicas mantidas em meio de cultura líquido sem reguladores de crescimento.

Synthetic seed technologies are useful tools for the field delivery of in vitro derived plantlets. In the present study, different encapsulation procedures and their efficacy in the plantlet regeneration using microshoots of banana cv. 'Grand Naine' were evaluated. Two encapsulation systems were evaluated: i) single encapsulation in beads or droplet hardening method; and ii) double layer or hollow beads. The use of different compounds to enhance the capsule conservation and the conversion to plantlets was also evaluated. The conversion capacity was assessed in vitro on water-agar culture medium or in ex vitro conditions with Gerbox® boxes. The single encapsulation system showed 80% conversion. The capsules with MS saline formulation treated with 100mM KNO3 showed 76% conversion. Capsules with 1.5g L-1 activated charcoal, and 0.5g L-1 benomyl sucrose-free capsules showed 75% conversion. The encapsulated and non-encapsulated microshoots exhibited 100% germination in response to MS culture medium, and polyethylene glycol after 10 days of storage at 4°C. Sucrose-free capsules showed significantly higher germination (83.3%) than those sucrose-enriched capsules (56.7%). The ex vitro conversion of encapsulated microshoots was 20% in the Gerbox™. These results indicate the feasibility using synthetic seeds in the large-scale micropropagation of banana cv. 'Grand Naine'.As tecnologias de semente sintética são ferramentas promissoras para a micropropagação de plantas. O objetivo do presente trabalho foi desenvolver um protocolo de encapsulamento e armazenamento de unidades encapsuláveis a partir de microbrotos de bananeira da cultivar 'Grand Naine'. Foram avaliados dois sistemas de encapsulamento: i) formação de cápsula pelo método de camada simples e ii) camada dupla. Foi avaliada também a adição de diferentes adjuvantes na matriz de alginato para promover a conservação da cápsula e favorecer a conversão em plantas. Para a conversão em planta foram avaliadas as condições in vitro sobre o meio de cultura composto de agar-água e as condições ex vitro em caixas Gerbox®. O melhor sistema de encapsulamento para os microbrotos da bananeira da cultivar 'Grand Naine' foi o de camada simples, com 80% de conversão. O emprego da formulação salina MS na forma de endosperma artificial e a descomplexação com 100mM KNO3 favoreceram a conversão de microbrotos (76%). A adição de carvão ativado (1,5g L-1) e de benomyl (0.5g L-1), na ausência de sacarose na matriz de alginato, resultou em 75% de conversão, além de reduzir a oxidação e a contaminação dos microbrotos. Microbrotos encapsulados e não-encapsulados revelaram 100% de germinação em resposta ao meio de cultura MS e PEG após 10 dias de estocagem a 4°C. Cápsulas sem a adição de sacarose resultaram em 83,3% de conversão, valor superior ao valor obtido com a adição de sacarose (56,7%). A conversão de microbrotos encapsulados foi de 20% nas condições ex vitro. Os resultados indicam a viabilidade de uso destas tecnologias para a micropropagação em larga escala da cultivar de banana 'Grand Naine'.

The Atlantic Forest is a biome with megadiversity and a number of bromeliads take part of it. This is the case of Vriesea reitzii, an endemic bromeliad threatened with extinction. Tissue culture techniques are valuable tools for the mass propagation of bromeliads, thus reducing pressure in the natural habitat. The aim of the present work was to establish an in vitro protocol based on the induction and proliferation of nodule cluster cultures of this species. Plantlets maintained in MS liquid culture medium plus NAA (2µM) and BAP (4µM) had the basal regions of leaves excised and then inoculated in gelled with agar (7g L-1) MS culture medium plus with Dicamba (2.5; 5; 10; 20 e 30µM) and Kin (2µM) or free of plant growth regulators. Nodule cluster cultures arose from the basal region of explants. The subculture to MS liquid medium plus GA3 (10µM) and in MS liquid medium free of plant growth regulators resulted in a high proliferation rate. The mean regenerative rate was 39 plantlets/0.03g of nodule culture. Plantlets were acclimatized in a mix substrate of 1:1 (v:v) of carbonized rice coat and Turfa Fertil® mineral supplement.A floresta tropical atlântica é um bioma de alta diversidade, rico em espécies endêmicas de bromélias, entre elas Vriesea reitzii, que se encontra ameaçada de extinção. Técnicas de cultura de tecidos possibilitam a propagação massal de bromélias, reduzindo a pressão de coleta na natureza. O presente trabalho teve como objetivo desenvolver um protocolo regenerativo baseado na indução e no desenvolvimento de culturas nodulares desta espécie. Plantas multiplicadas em meio MS líquido com ANA (2µM) e BAP (4µM) tiveram suas folhas excisadas e inoculadas em meio MS geleificado com ágar (7g L-1) e suplementado com Dicamba (2,5; 5; 10; 20 e 30µM) e Kin (2µM) ou isento de fitorreguladores. A indução de culturas nodulares foi observada na região basal dos explantes. Estas culturas foram subcultivadas em meio de cultura MS líquido suplementado com GA3 (10µM) e subseqüentemente para meio MS líquido isento de fitorreguladores, resultando em altas taxas de proliferação de culturas nodulares que originaram brotos adventícios e microbrotos. A taxa média de regeneração foi de 39 brotos/0,03g de culturas nodulares. As mudas foram aclimatizadas com sucesso em substrato composto por fertilizante organomineral Turfa Fértil® e casca de arroz carbonizada na proporção de 1:1 (v:v).

The objective of this work was to characterize the grape germplasm in Santa Catarina, Brazil, using microsatellite DNA markers (simple sequence repeats - SSR). The DNA samples were collected from leaves and shoots of accessions of public and private collections from the counties Urussanga, Nova Trento, Rodeio, São Joaquim, Campos Novos, Videira, and Água Doce. Ten SSR loci (VVS2, VVMD5, VVMD7, VVMD27, VrZAG62, VrZAG79, VVMD25, VVMD28, VVMD31, and VVMD32) were analysed by capillary electrophoresis. Molecular profiling was conducted for 190 grapevines (European, American, and hybrids), and 67 genotypes were obtained. The data were compared with each other and with those from the literature and from online databases, in order to identify varieties and discover cases of synonymy and homonymy. Forty molecular profiles corresponded to known varieties, while 27 genotypes were described for the first time. The existence of typical germplasm composed mainly of American and hybrid varieties is an important finding for local viticulture. Applications of the results rely on quality control and certification at the nursery level. Increasing precision in the characterization of grapevine genotypes may help breeding programs.O objetivo deste trabalho foi caracterizar a diversidade de videiras em Santa Catarina, por meio de marcadores moleculares microssatélites ("simple sequence repeats" - SSR). Amostras de DNA foram coletadas a partir de folhas e ramos de acessos de coleções de germoplasma públicas e privadas, nos municípios de Urussanga, Nova Trento, Rodeio, São Joaquim, Campos Novos, Videira e Água Doce. Dez loci SSR (VVS2, VVMD5, VVMD7, VVMD27, VrZAG62, VrZAG79, VVMD25, VVMD28, VVMD31 e VVMD32) foram analisados por eletroforese capilar. Foram produzidos perfis moleculares de 190 acessos de videira (europeus, americanos e híbridos), e 67 genótipos foram individualizados. Os dados foram comparados entre si e com aqueles disponíveis em literatura e em bancos de dados online, para a identificação de correspondências e casos de sinonímia e homonímia. Quarenta perfis moleculares corresponderam a variedades conhecidas, e 27 genótipos foram descritos pela primeira vez. A existência de um germoplasma típico, composto principalmente de variedades americanas e híbridas, é um fator importante para a viticultura local. A aplicação desses resultados poderá contribuir para o controle de qualidade e a certificação de mudas. Além disso, aumentar a precisão no que tange à caracterização genética da videira, auxiliará os programas de melhoramento genético.

The aim of this work was to quantify the protein, starch and total sugars levels during histodifferentiation and development of somatic embryos of Acca sellowiana Berg. For histological observations, the samples were dehydrated in a battery of ethanol, embedded in historesin and stained with toluidine blue (morphology), coomassie blue (protein bodies) and periodic acid-Schiff (starch). Proteins were extracted using a buffer solution, precipitated using ethanol and quantified using the Bradford reagent. Total sugars were extracted using a methanol-chloroform-water (12:5:3) solution and quantified by a reaction with anthrone at 0.2%. Starch was extracted using a 30% perchloric acid solution and quantified by a reaction with anthrone at 0.2%. During the somatic embryogenesis' in vitro morphogenesis and differentiation processes, the total protein levels decreased and the soluble sugars levels increased during the first 30 days in culture and remained stable until the 120th day. On the other hand, total protein levels increased according to the progression in the developmental stages of the somatic embryos. The levels of total sugars and starch increased in the heart and cotyledonary stages, and decreased in the torpedo and pre-cotyledonary stages. These compounds play a central role in the development of somatic embryos of Acca sellowiana.O objetivo deste trabalho foi quantificar os teores de proteína, amido e açúcares totais durante a histodiferenciação e desenvolvimento dos embriões somáticos em Acca sellowiana Berg. Para as observações histológicas, as amostras foram desidratadas em uma bateria de etanol, emblocadas em historesina e coradas com azul de toluidina (morfologia), azul de coomassie (corpos proteicos) e reativo ácido periódico de Schiff (amido). As proteínas foram extraídas usando uma solução tampão, precipitadas usando etanol e quantificadas por meio do reativo de Bradford. Os açúcares totais foram extraídos usando uma solução metanol-clorofórmioágua (12:5:3) e quantificados pela reação com antrona a 0,2%. O amido foi extraído usando uma solução de ácido perclórico a 30% e quantificado pela reação com antrona a 0,2%. Durante a diferenciação e morfogênese in vitro da embriogênese somática, os teores de proteínas totais decresceram e os açúcares solúveis aumentaram durante os 30 primeiros dias em cultura e permaneceram constantes até os 120 dias. Por outro lado, os teores das proteínas totais apresentaram incremento de acordo com a progressão nos estádios de desenvolvimento dos embriões somáticos. Os teores de açúcares totais e de amido aumentaram nos estádios cordiforme e cotiledonar e diminuíram nos estádios torpedo e pré-cotiledonar. Esses compostos exercem papel central no desenvolvimento de embriões somáticos de Acca sellowiana.

A Acca sellowiana Berg. (Myrtaceae) é uma frutífera nativa dos planaltos meridionais do Sul do Brasil e que se encontra em processo de domesticação. Seus frutos são doce-acidulados e podem ser consumidos in natura ou empregados para a produção de sucos e doces. Assim, informações sobre o desenvolvimento, morfologia e anatomia de seus frutos são de grande interesse e foram objetos do presente trabalho. O fruto (ovário mais hipanto), no tempo zero (plena floração), tem, em média, 0,6 cm de altura e 0,4 cm de diâmetro, sendo cerca de dez vezes menor que o fruto maduro. Longitudinalmente, identificam-se três regiões distintas: locular, sublocular e prolongamento. Transversalmente, na região mediana, estão delimitadas três regiões: 1) epiderme (casca): com tricomas unicelulares e simples; 2) região parenquimática: rica em braquiesclereídes, isoladas ou em pequenos grupos (2-3 células), com oito feixes vasculares concêntricos perifloemáticos distribuídos radialmente e muitas glândulas esféricas subepidérmicas; 3) região interna (polpa): com células pequenas, cúbicas, nitidamente dispostas em 3-4 camadas ao redor dos lóculos, várias contendo drusa. Quatro lóculos são separados pelos septos e vários óvulos nascem de placentas axiais, com duas fileiras por lóculo. Não ocorrem nectários. À medida que o fruto se desenvolve, surgem, na região intermediária, grupos de células de paredes finas, que crescem muito e diferenciam-se em braquiesclereídes. As placentas crescem, ocupando todo o espaço interior dos lóculos à medida que estes aumentam de tamanho e as sementes se desenvolvem. Assim, o fruto maduro apresenta uma região periférica de consistência firme e gosto adstringente, e uma região central macia e adocicada.Acca sellowiana Berg. (Myrtaceae) is a fruit-bearing treelet or shrub native from the highlands of South Brazil. The plant is currently in the domestication process. Its fruit is sweet-acidified and can be consumed raw or be used for the preparation of juice and jam. Thus, information on the development, morphology and anatomy of the fruit is of great interest, and was featured as the objective of this study. The average fruit dimensions (ovary surrounded by the hypanthium) in the bloom stage were 0.6 cm in length and 0.4 cm in diameter, being ten times smaller than the ripe fruit. Longitudinally, three distinct regions were observed: a locular-, a sublocular- and a prolongation region. In a transverse cross-section in the middle of the fruit, three regions were delimited: 1) epidermis (peel) with simple unicellular trichomes. 2) parenchymatous region (flesh) rich in stone cells, isolated or aggregated in small groups of 2-3 cells; with eight radially distributed concentric periphloematic vascular bundles; and with many spherical glands near the epidermis. 3) inner region (pulp) with small cubic cells, organized in 3-4 layers around the locules, various containing druses. The four locules are separated by septa. The numerous ovules originate from axillary placentas, with two rows per locule. No nectaries were observed. As the fruit develops, groups of thin-walled cells appear in the intermediate region. These cells become very large and transform into stone cells. The placentas grow and occupy the whole space within the growing locules, as these locules grow bigger and seeds develop. Thus, the ripee fruit has a peripheral region of firm consistence and astringent taste, and a soft sweet core.

Acca sellowiana (Myrtaceae) é uma frutífera nativa da região Sul do Brasil e nordeste do Uruguai, que vem despertando grande interesse devido ao alto potencial organoléptico de seus frutos. Neste trabalho, teve-se como objetivo a caracterização do tipo de sistema de incompatibilidade atuante em A. sellowiana, através da avaliação do desenvolvimento dos tubos polínicos. Utilizaram-se dois acessos: 458, sendo autocompatível, e 101, auto-incompatível. A maior porcentagem de germinação de grãos de pólen foi observada no acesso 101, com 68,8% de grãos de pólen germinados. Não foram observadas diferenças no crescimento dos tubos polínicos em pistilos autopolinizados ou de polinização cruzada, em ambos os acessos. O crescimento completo do tubo polínico até o ovário ocorreu em pistilos coletados 96 horas após a polinização, independentemente do tratamento aplicado. Sugere-se a ocorrência de auto-esterilidade ou auto-incompatibilidade tardia ou pós-zigótica, considerando que o abortamento dos frutos de A. sellowiana é uniforme e ocorre num prazo curto, por volta de 20 a 30 dias após a fertilização das flores.Acca sellowiana (Myrtaceae) is a native fruit species from the South Brazil and northeast Uruguay which is attracting great interest due to the high organoleptic potential of its fruits. In this study we aimed to characterize the type of incompatibility system which occurs in A. sellowiana, through the evaluation of the polinic tube development. Two accesses were used: being 458 self-compatible and 101 self- incompatible. The highest percentage of pollen grains germination, 68,8%, was observed in the access 101. It was not observed differences in the growth of the polinic tubes between self-pollinated pistils and cross-pollinated ones in both accesses. The complete growth of the polinic tube until reaching the ovary occurred only in pistils collected 96 hours after the polinization, independent of the treatment. The data suggests the occurrence of self-sterility, or a delayed or post-zygotic self-incompatibility in the species, considering that the abortion of A. sellowiana fruits is uniform and occurs in a short period of time, around 20 to 30 days, after the fertilization of the flowers.

The pineapple guava (Acca sellowiana), known in portuguese as the goiabeira-serrana or "Feijoa", is a native fruit tree from southern Brazil and northern Uruguay that has commercial potential due to the quality and unique flavor of its fruits. Knowledge of genetic variability is an important tool in various steps of a breeding program, which can be facilitated by the use of molecular markers. The conservation of repeated sequences among related species permits the transferability of microsatellite markers from Eucalyptus spp. to A. sellowiana for testing. We used primers developed for Eucalyptus to characterize A. sellowiana accessions. Out of 404 primers tested, 180 amplified visible products and 38 were polymorphic. A total of 48 alleles were detected with ten Eucalyptus primer pairs against DNA from 119 A. sellowiana accessions. The mean expected heterozygosity among accessions was 0.64 and the mean observed heterozygosity 0.55. A high level of genetic diversity was also observed in the dendrogram, where the degree of genetic dissimilarity ranged from 0 to 65% among the 119 genotypes tested. This study demonstrates the possibility of transferring microsatellite markers between species of different genera in addition to evaluating the extent of genetic variability among plant accessions.

Passiflora cincinnata Mast. é uma espécie silvestre de maracujazeiro de ampla distribuição. Possui hábito trepador e crescimento vigoroso, com flores muito vistosas e perfumadas. O trabalho teve como objetivo a produção de sementes sintéticas da espécie P. cincinnata, utilizando-se de embriões somáticos e zigóticos em diferentes formas de cultivo. Os embriões somáticos em estágio de desenvolvimento pré cotiledonar e cotiledonar foram obtidos a partir de embriões zigóticos cultivados em meio de MS na presença de 18,1 μM de 2,4-Ácido-dichlorophenoxiacetico (2,4-D) e 4,5 μM Benziladenine (BA). Embriões zigóticos e embriões somáticos foram encapsulados em matriz de alginato de sódio a 2,5% (p v-1) e complexados em solução estéril de CaCl2.2H2O a 0,1M. Embriões somáticos e zigóticos foram encapsulados na Matriz (I) com alginato de sódio, Matriz (II) com alginato de sódio + endosperma artificial e Matriz (III) - alginato de sódio + endosperma artificial e suplementado com 0,15% (p v-1) de carvão ativado. Embriões zigóticos cultivados em frascos encapsulados na Matriz (I) 79% germinaram, 76% Matriz (II) e na Matriz (III) 86%. Os embriões somáticos cotiledonares encapsulados nos três tipos de matrizes responderam com maior percentual de germinação quando cultivados em plugs de celulose com mais de 50% de embriões convertidos. Os embriões somáticos pre cotiledonares encapsulados nos três tipos de matrizes e nas diferentes formas de cultivo não responderam. No cultivo ex vitro nos dois tipos de substratos PlantMax e Florialite o número de embriões convertidos foi baixo, sendo o melhor resultado com 12,67 % no Florialite e encapsulados na Matriz (I).

During the evolution of the eukaryotic cell, plastids and mitochondria arose from an endosymbiotic process, which determined the presence of three genetic compartments into the incipient plant cell. After that, these three genetic materials from host and symbiont suffered several rearrangements, bringing on a complex interaction between nuclear and organellar gene products. Nowadays, plastids harbor a small genome with ~130 genes in a 100-220 kb sequence in higher plants. Plastid genes are mostly highly conserved between plant species, being useful for phylogenetic analysis in higher taxa. However, intergenic spacers have a relatively higher mutation rate and are important markers to study genetic diversity and divergence within natural plant populations. The predominant uniparental inheritance of plastids is like a highly desirable feature for phylogeny studies. Moreover, the gene content and genome rearrangements are efficient tools to capture and understand evolutionary events between different plant species. Currently, genetic engineering of the plastid genome (plastome) offers a number of attractive advantages as high-level of foreign protein expression, marker-gene excision, gene expression in operon and transgene containment because of maternal inheritance of plastid genome in most crops. Therefore, plastid genome can be used for adding new characteristics related to synthesis of metabolic compounds, biopharmaceutical and tolerance to biotic and abiotic stresses. Here, we describe the importance and applications of plastid genome as tools for genetic and evolutionary studies, and plastid transformation focusing on increasing the performance of horticultural species in the field.

The objetive of this work was to rescue immature embryos of apple rootstocks Malus prunifolia (Marubakaido) and Malus pumila (M9) after 40-60 days of pollination and to put them into MS culture media supplemented with agar (6 g L-1) and casein hydrolysate (500 mg L-1). Embryos originated from interspecific crosses and open pollination showed differences in the in vitro responses, depending on the female parent, the developmental stage of the embryo, and the culture medium composition. Embryos of the M. pumila rootstock, rescued within 40 days after pollination and put in culture medium supplemented with indolacetic acid (IAA), gibberellic acid (GA3), kinetin and maltose, resulted in a normal development of plantlets. However, embryos originating from hand-pollination, cultivated in medium supplemented with 14 µM IAA, 5 µM kinetin and 1.5 µM Ga3 (MS1), mainly those of M. prunifolia x M. pumila, showed a high percentage of rusted embryos (96.2%). Embryos from open pollination of M. prunifolia and M. pumila formed calluses. It was possible to identify the influence of the female parent by the enhanced development of M. pumila shoots derived from open or hand-pollination. The crossing of responsive species and the use of the technique of embryo culture provided a rapid and uniform germination and, consequently, the development of fully normal seedlings.O objetivo deste trabalho foi resgatar embriões imaturos de porta-enxertos de macieira Malus prunifolia (Marubakaido) e Malus pumila (M9) depois de 40 e 60 dias de polinização e colocá-los em meio de cultura MS suplementado com ágar (6 g L-1) e hidrolisado de caseína (500 mg L-1). Embriões originados do cruzamento interespecífico dirigido e de polinização aberta mostraram diferenças significativas in vitro, tendo sido observadas diferenças quanto ao progenitor feminino, quanto à fase de desenvolvimento do embrião e à composição do meio de cultura. Embriões do porta-enxerto M. pumila, resgatados aos 40 dias depois da polinização e colocados em meio de cultura suplementado com ácido indolacético (AIA), ácido giberélico (GA3), cinetina e maltose, resultaram em desenvolvimento normal das plantas. Porém, foi observada alta porcentagem de embriões oxidados (96,2%), originados de polinização dirigida, cultivados em meio com 14 µM de AIA, 5 µM de cinetina e 1,5 µM de GA3 (MS1), principalmente para o cruzamento M. prunifolia e M. pumila. Embriões de polinização aberta de M. prunifolia x M. pumila formaram calos. Foi possível identificar a influência do progenitor feminino, pelo incremento de brotações de M. pumila derivada de polinização aberta e dirigida. O cruzamento de espécies responsivas e o uso da técnica de cultura de embrião promoveram germinação rápida e uniforme e, por conseguinte, o desenvolvimento de mudas normais.

Different factors associated to somatic embryogenesis induction in Euterpe edulis were investigated in thepresent study. Zygotic embryos were inoculated in MS culture medium supplemented with 2.4-D (0, 30, 35, 40 mg L-1), 3 mgL-1 2iP, 0.5 g L-1 glutamine, 0.5 g L-1 activated charcoal; 30 g L-1 glucose or sucrose and gelled with 5 g L-1 Agar. Leaf sheathswere inoculated on MS culture medium supplemented with Picloram (72.3 mg L-1) or 2.4-D (66.3 mg L-1), 3 mg L-1 2iP,glutamine (0; 0.29; 0.58; 1.17 g L-1), 1.5 g L-1 activated charcoal and gelled with 2.5 g L-1 Phytagel®. Indirect somatic embryoswere induced from zygotic embryos on culture medium with 40 mg L-1 2.4-D and 30 g L-1 sucrose. The highest percentage ofcallus formation on leaf sheaths occurred in MS culture medium supplemented with 1.17 g L-1 glutamine and 66.3 mg L-1 2.4-D.

No presente trabalho, estudaram-se características associadas à germinação in vitro e ao desenvolvimento in vivo do tubo polínico em seis variedades-copa e de porta-enxertos de macieira como subsídios para o estabelecimento de programas de melhoramento genético. Utilizou-se de pólen de seis cultivares de macieira inoculado em meio de cultura com ágar (10 g.L-1) em água destilada, combinados com concentrações de sacarose (0; 10; 20; 30; 40 e 50%) e ácido bórico (0 e 40 mg.L-1). Para o estudo do desenvolvimento do tubo polínico, realizou-se coleta das flores em quatro períodos (6; 12; 24 e 48 horas após as polinizações) em M9 x Marubakaido e a autofecundação em M9, sendo os tubos polínicos analisados em coloração de azul de anilina acidificada/carmim acético e em fluorescência. A sacarose, em concentrações entre 15% a 25%, pode ser empregada com sucesso para a germinação in vitro de grãos de pólen da macieira. O ácido bórico não teve efeito positivo para esta característica. Na ausência do ácido bórico e na presença de 15% de sacarose, observaram-se os maiores percentuais de germinação: Fuji (51,1%), Imperatriz (31,7%), M.9 (20,8%), Catarina (19,2%), Gala (13,7%) e Marubakaido (6,1%). Quanto ao desenvolvimento do tubo polínico, com 12 horas da polinização, iniciou-se a germinação no pólen, no estigma, no cruzamento M.9 x Marubakaido, e após 24 horas da polinização observou-se 83% de germinação. As técnicas de coloração com azul de anilina acidificada com carmim acético e de visualização em fluorescência foram eficientes na visualização e coloração dos grãos de pólen e do desenvolvimento dos tubos polínicos.In the present work it was studied features associated to the germination of pollen and the development of pollen tubes in six apple varieties as subsidies for the establishment of genetic improvement programs. Pollen grains were inoculated in culture medium with distilled water, agar (10 g. L-1), combined with sucrose at 0, 10, 20, 30, 40 and 50% and boric acid at 0 and 40 mg. L-1. For the study of pollen tubes development flowers were collected at 6, 12, 24 and 48 hours after antesis in M.9 x Marubakaido and in the self-pollinated M.9, being the pollen tubes analyzed in staining with blue aniline with acetic carmim and in fluorescence. Sucrose in concentrations ranging from 15% to 25% can successfully be used for the in vitro germination of pollen grains. Boric acid was not effective in promoting in vitro germination of pollen grains. In the absence of boric acid and in the presence of 15% of sucrose it was observed the largest germination rates: Fuji (51.1%), Imperatriz (31.7%), M.9 (20.8%), Catarina (19.2%), Gala (13.7%), and Marubakaido (6.1%). In the crossing M.9 x Marubakaido after 12 hours of pollination the pollen germination began in the stigma and after 24hrs it was observed 83% of germination. The techniques employed were efficient in the visualization and coloration of the pollen grains and of development of tube pollen.

A goiabeira-serrana (Acca sellowiana Berg.) é uma mirtácea nativa do planalto meridional brasileiro, com dispersão secundária no Uruguai, e produz um fruto de sabor único. Plantas de um pomar comercial composto por duas famílias de meios-irmãos (FMI1 e FMI2) foram avaliadas, com o objetivo principal de analisar a variabilidade fenotípica de várias características. As médias (± desvio-padrão) obtidas para as características avaliadas, no ano de 2000, foram: produtividade (29,2±35,0 e 55,1±53,4 frutos por planta); altura (2,2±0,7 m e 2,7±0,8 m); diâmetro de copa (1,9±0,7 m e 2,1±0,7 m); número de ramificações a 20 cm do solo (2,4±1,5 e 1,6±1,2) e distância entre o estigma e as anteras na flor (0,5±0,2 cm e 0,2±0,2 cm), para as plantas da FMI1 e da FMI2, respectivamente. As avaliações revelaram diferenças estatisticamente significativas entre as duas famílias para as características avaliadas, com exceção de diâmetro de copa. As correlações entre características vegetativas não foram estatisticamente significativas, com exceção da correlação entre a altura e a distância entre estigma e anteras (r=-0,53) para a FMI1, no ano de 2000. As regressões não foram significativas, com exceção da regressão entre altura e produtividade na FMI1, em 1998, e entre a produtividade e a distância entre estigma e anteras na FMI2, em 2000. Porém, todas apresentaram coeficientes de determinação inferiores a 0,15.Feijoa (Acca sellowiana Berg.) is a native Myrtaceae of the Brazilian Southern plateau with secondary dispersion in Uruguay and its fruit has a unique flavor. Plants of a commercial orchard of this species originated by two half-sib families (FMI1 and FMI2) were evaluated, with the objective to characterize the phenotypic variation of several traits. The means (±standard deviation) of the evaluated traits, obtained in 2000 were: productivity (29.2±35.0 and 55.1±53.4 fruits per plant); height (2.2±0.7 m and 2.7±0.8 m), canopy diameter (1.9±0.7 m, and 2.1±0.7 m), number of ramifications at 20 cm of the soil (2.4±1.5 and 1.6±1.2) and distance between the stigma and the anthers in the flower (0.5±0.2 cm and 0.2±0.2 cm) for FMI1 and FMI2, respectively. The evaluations revealed significant statistical differences between the two families for plants traits evaluated, except for canopy diameter. The correlations among vegetative traits were not significant, except for height x distance between stigma and anthers (r=-0.53) for FMI1 in 2000. The regressions were not significant, except the regressions between height and productivity for FMI1 in 1998 and between productivity and distance between stigma and anthers in FMI2, in 2000, all with R²

The aim of the present work was to establish in vitro conditions for the induction, stabilization and proliferation of embryogenic cultures of A. angustifolia. Pre-cotyledonary staged zygotic embryos inoculated BM medium supplemented with 5 µM 2,4- D, 2 µM BAP and Kin, and 3% maltose or sucrose resulted in 66.7% induction rate. The rate of induction of embryogenic cultures was affected by the carbon source, as well the multiplication and morphology of the embryogenic cultures. Embryogenic cultures maintained in BM medium with maltose presented bipolar morphology. Globular somatic embryos were obtained BM medium with 9% (PEG) and (9%) maltose. These results could establish an in vitro regenerative protocol towards the conservation and improvement of this important species.No presente trabalho foram investigadas as condições para a indução, estabilização e proliferação de culturas embriogênicas de A. angustifolia. Foram também descritos os padrões de morfogênese e histodiferenciação de culturas embriogênicas e embriões somáticos desta espécie em resposta a diferentes fontes e concentrações de carboidratos. Embriões zigóticos no estágio pré-cotiledonar, inoculados em meio de cultura BM, suplementados com 5 µM de 2,4-D, 2 µM de BAP e Kin e 3% de maltose ou sacarose resultaram em uma taxa de indução de 66,7%. A fonte de carbono afetou a taxa de indução, a multiplicação e a morfologia das culturas. Culturas embriogênicas mantidas em meio de cultura BM suplementado com maltose apresentaram morfologia bipolar. Embriões somáticos globulares foram obtidos em meio de cultura BM suplementado com PEG e maltose. Os resultados obtidos permitirão avançar na direção de um protocolo regenerativo in vitro visando à conservação e o melhoramento genético desta espécie.

O presente trabalho teve como objetivo o estabelecimento de um protocolo de regeneração in vitro de mudas de Aspidosperma polyneuron (peroba-rosa), a partir de segmentos nodais de material juvenil. Brotações apicais de mudas de dois anos de idade foram desinfestadas com 0,25% de hipoclorito de sódio ou 0,05% de cloreto de mercúrio, durante 10 min, visando ao estabelecimento de culturas assépticas. A indução de brotações múltiplas foi realizada em meio de cultura WPM, suplementado com BAP, ZEA ou CIN (2,2-8,8 miM), no cultivo inicial e nos dois subcultivos subseqüentes. Para indução de brotações alongadas foram testadas as combinações de fitorreguladores: 2,25 miM de BAP, ZEA ou CIN, associadas com 1,25 miM de AIB. A indução de raízes foi avaliada com tratamentos em soluções de AIB (2,5-10 mM), durante 5 ou 15 min. As mudas enraizadas foram transplantadas para casa de vegetação. A desinfestação das brotações apicais foi eficiente com 0,25% de NaOCl ou 0,05% de HgCl2, durante 10 min, obtendo-se 72,89 e 84,10% de sobrevivência, respectivamente. As maiores taxas médias de regeneração de brotações axilares (4 a 5) foram obtidas em meio de cultura suplementado com ZEA ou BAP (4,4-8,8 miM), após o segundo subcultivo. Concentrações mais reduzidas de BAP ou ZEA (2,25 miM) e 1,25 miM de AIB proporcionaram, em média, três brotações mais alongadas (1,5-2,5 cm de comprimento). Tratamentos com soluções de 10 mM de AIB, durante 15 min, foram eficientes na indução de raízes (80%), e as mudas transplantadas apresentaram taxas de sobrevivência superiores a 90% em casa de vegetação.The objective of the present work was to establish a micropropagation protocol of Aspidosperma polyneuron from juvenile material. Apical shoots from two years old seedlings were collected in a greenhouse and sterilised with NaOCl or HgCl2 to establish aseptic cultures. Multiple shoots induction was evaluated in WPM medium, supplemented with BAP, ZEA or KIN (2.2 - 8.8 muM) in initial culture and two subsequent subcultures. The elongation of shoots was tested with growth regulators combinations: 2.25 muM of BAP, ZEA or KIN with 1.25 muM of IBA. IBA treatments (2.5; 5.0 and 10 mM) were tested with 5 and 15 minutes to induce roots. Plantlets were planted in a greenhouse. Efficient apical shoots sterilization was achieved with NaOCl (0,25% - 10 minutes) or HgCl2 (0.05%-10 minutes); survival rates were 72.89% and 84,10%, respectively. Apical shoots induced 4-5 axillary buds in WPM culture medium, containing ZEA or BAP (4.4 - 8.8 muM) following two subcultures. Reduced concentrations of ZEA or BAP (2.25 muM), combined with IBA (1.25) produced elongated shoots. IBA treatment (10 mM) during 15 minutes induced higher rooting percentages (80%). Plantlets planted in a greenhouse showed higher survival rates (90%).

A method for the isolation of coupled mitochondria from the callus of Araucaria angustifolia is described for the first time. Mitochondria were isolated from embryogenic callus of A. angustifolia. They were metabolically active, able to sustain oxidative phosphorylation as shown by respiratory control ratio values, which were about 2.4 when respiring on succinate as substrate. Oxygen uptake experiments, using freeze-thawed disrupted mitochondria, showed the presence of alternative rotenone-insensitive NAD(P)H dehydrogenases, which were stimulated by Ca2+. The procedure now described for the isolation of A. angustifolia mitochondria is an important new tool, allowing the investigation of mitochondrial bioenergetics and metabolism and physiology of plants.Um procedimento de isolamento de mitocôndrias funcionalmente intactas de calos embriogênicos de Araucaria angustifolia foi desenvolvido pela primeira vez em nosso laboratório. Mitocôndrias isoladas por este método são metabolicamente ativas, capazes de sustentar fosforilação oxidativa como mostrado pelo controle respiratório de aproximadamente 2,4, respirando na presença de succinato como substrato. Através de experimentos de consumo de oxigênio com mitocôndrias rompidas em nitrogênio líquido foi demonstrada a presença de NAD(P)H desidrogenases alternativas, insensíveis à rotenona e estimuladas por Ca2+. O isolamento de mitocôndrias de A. angustifolia é um novo e importante instrumento para estudar plantas, permitindo a execução de múltiplas investigações a respeito da bioenergética mitocondrial e fisiologia vegetal.

This study was aimed at adapting a DNA extraction protocol by Araucaria angustifolia leaves, and testing the informative capacity of RAPD markers for genetics diversity analysis in natural populations of this species. The extraction method was standardized by eight tested protocols and it was possible to obtain good quality DNA for RAPD reactions. The OD260/OD280 ratio ranged from 1.7 to 2.0 in 80% of the samples, indicating that they had a low level of protein contamination. The RAPD markers demonstrated their capacity as a useful tool to genetic studies, since the estimate of a genetic diversity (Shannon index H=0.50) higher than that obtained by other authors with isozyme markers (H=0.29) and PCR-RFLP (H=0.0) for the same natural population of the Ecological Municipal Park, Lages, Santa Catarina, Brazil.

Frutos de um pomar comercial composto por progênies de duas plantas matrizes foram avaliados, com o objetivo de caracterizar a variabilidade fenotípica. As características peso de fruto, diâmetro e sólidos solúveis totais apresentaram diferenças estatísticas significativas entre as médias de famílias em pelo menos dois dos anos avaliados, enquanto para comprimento e rendimento de polpa a diferença não foi significativa. Houve diferença significativa entre médias de anos, dentro de cada família, para todas as características, com exceção das médias de peso de fruto entre 1998 e 1999 nas duas famílias e as médias de sólidos solúveis totais entre 1998 e 1999 em uma família e entre 1999 e 2000 na outra família. As correlações e regressões entre características produtivas mais relevantes foram obtidas entre peso de fruto e peso de casca, peso de fruto e comprimento, peso de fruto e diâmetro, peso de casca e comprimento, peso de casca e diâmetro e comprimento e diâmetro.Fruits of a commercial orchard originated by seedlings from two plants were evaluated, with the objective to characterize the phenotypic variation. Statistical differences were found among family means in at least two years for fruit diameter and solids soluble contents, but not for fruit length and pulp revenue. There were statistically significant differences in means among year means, within each family, for all the fruits traits, except for fruit weight between 1998 and 1999 in both families, and for soluble solids contents between 1998 and 1999 for family 1 and between 1999 and 2000 for the other family. The most important correlations and regressions among productive traits were obtained among fruit weight and peel weight, fruit weight and length, fruit weight and diameter, peel weight and length, peel weight and diameter and length and diameter.