Estabilidad genética entre clones de berenjena in vitro inducidos por diferentes reguladores de crecimiento de plantas

Many factors may influence the genetic stability of plant in vitro clones, among which the genotype and the regenerative process induced by plant growth regulators. The resulting somaclonal variations may be useful for breeding projects, but may be detrimental to germplasm conservation. The objective of this work was to evaluate the genetic stability of Solanum melongena cv. Florida Market clones, obtained in response to different plant growth regulators. For the production of clones, leaf explants were used from commercial seed germinated plants. The explants were inoculated in Murashige and Skoog medium supplemented with different plant growth regulators at pre­defined concentrations. The DNA was extracted by the CTAB method from leaves of complete plants obtained by somatic embryogenesis induced by naphthaleneacetic acid (NAA) or indirect organogenesis induced by benzylaminopurine (BAP) or thidiazuron (TDZ). For the RAPD, 117 DNA samples were amplified by ten decamer primers and 49 specific bands were selected among the products for the comparative study. A total of 5733 fragments were obtained, with a rate of 5.37% polymorphism. NAA did not generate polymorphism and the BAP was responsible for the highest rate obtained (14.28%). Two RAPD primers were identified as markers for monitoring the genetic stability of eggplant. The polymorphic pattern was observed only in clones originating from indirect organogenesis. These results indicate the usefulness of a monitoring protocol for studies using in vitro cloned eggplant.Muchos factores pueden influir en la estabilidad genética de los clones de plantas in vitro, entre los que se encuentran el genotipo y el proceso regenerativo inducido por los reguladores del crecimiento. Por lo tanto, las variaciones somaclonales resultantes del cultivo pueden ser útiles para proyectos de mejoramiento genético, pero pueden ser perjudiciales para la conservación de germoplasma. El objetivo de este estudio fue evaluar la estabilidad genética de Solanum melongena cv. Florida Market, obtenida en respuesta a diferentes reguladores del crecimiento. Para la producción de clones se utilizaron explantes de hojas provenientes de plantas obtenidas de semillas germinadas. Los explantes se inocularon en medio de cultivo Murashige y Skoog con los diferentes reguladores del crecimiento en concentraciones predefinidas. El ADN se extrajo mediante el método CTAB a partir de plantas completas obtenidas por medio de embriogénesis somática inducida por ácido naftalenoacético (ANA) u organogénesis indirecta inducida por bencilaminopurina (BAP) o tidiazurón (TDZ). Para el RAPD, 117 muestras de ADN se amplificaron mediante diez cebadores y se seleccionaron 49 bandas puntuales entre los productos, para el estudio comparativo. Se obtuvieron un total de 5733 fragmentos, con una tasa de 5.37% de polimorfismo. ANA no generó polimorfismo y BAP fue responsable de la tasa más alta obtenida (14.28%). Se identificaron dos cebadores RAPD como marcadores para monitorear la estabilidad genética de la berenjena. El patrón polimórfico se observó solo en los clones originados en la organogénesis indirecta. Estos resultados indican la utilidad de un protocolo de monitoreo para estudios que usan berenjena clonada in vitro. Palabras clave: Solanum melongena, cultivo in vitro, variación somaclonal, RAPD, polimorfismo de ADN, fitomejoramient

Evaluation of cryopreservation of Petiveria alliacea somatic embryos based on stress caused for the method used / Avaliação da criopreservação de embriões somáticos de Petiveria alliacea com base no estresse causado pelo método usado

Petiveria alliacea is a medicinal species with great potential for pharmacological use against several pathologies, including neoplasms. Many studies have been developed to optimize efficient production methodologies and long-term conservation of this species' genetic resources, with a view to phytochemical and pharmacological research. This study demonstrates the efficiency of the D-cryoplate technique, applied to somatic embryos from plants maintained in vitro. Leaf explants were inoculated in culture medium containing 20 ?M PIC and incubated under standard conditions in plant tissue culture. After 60 days, the somatic embryos induced directly on the leaf tissue surface were transferred to the multiplication medium (MS0). For cryopreservation, samples of these embryos were precultured for 24 hours in medium supplemented with sucrose (0.5M), then groups of 3-5 somatic embryos were encapsulated in calcium chloride directly in aluminum cryoplates. The cryoplates with somatic embryos adsorbed were immersed in a conditioning solution (loading) for 20 min. After being removed from loading, the somatic embryos adhered to the cryoplates were exposed to laminar flow air for different periods of time (0 to 140 min) to assess the level of dehydration. Then, samples submitted at each time were immersed in liquid nitrogen for 2 min. After this time, the cryoplates were removed and kept at room temperature for 20 min, and the somatic embryos were cultivated in MS0 medium. Evaluation after each treatment showed a high survival rate (93%) in cryopreserved somatic embryos. After 90 days of culture it was observed that somatic embryos dehydrated for 120 min showed the highest multiplication rate (32 embryos/inoculated embryo) obtained so far with these explants. The D-cryoplate technique brought innovation to established protocols representing the best option for in vitro conservation of these structures biotechnologically produced that are so promising for phytochemical and pharmacological research

CONSERVAÇÃO IN VITRO DA BIODIVERSIDADE VEGETAL: ABORDAGEM NO ENSINO MÉDIO

As plantas constituem a base dos ecossistemas e a necessidade de conservar a biodiversidade é imprescindível para a sociedade. O objetivo deste projeto foi conscientizar os alunos de uma escola pública de ensino médio do Rio de Janeiro desta realidade, gerando oportunidade para graduandos e pós-graduandos transmitirem conceitos de biotecnologia aplicada à conservação de plantas. A metodologia consistiu de palestras, documentários em vídeo e visita técnica aos laboratórios do Núcleo de Biotecnologia Vegetal da Universidade do Estado do Rio de Janeiro, onde os alunos acompanharam a produção e a conservação de plantas in vitro. A avaliação baseou-se na frequência e na participação, além da comparação das respostas a um questionário aplicado no início e ao final do projeto. Foi detectada uma grande variação entre as turmas, tanto na frequência (41-68%), quanto no percentual de acertos nos questionários, inicial (42-57%) e final (53-68%). Contudo, todas as turmas avaliadas apresentaram maior rendimento após a participação no projeto, indicando um efeito positivo nas questões específicas de Biotecnologia e sua aplicação na conservação da Biodiversidade. Palavras-chave: Desenvolvimento Sustentável, Cultura In Vitro, Ensino de Biologia, Biotecnologia Vegetal   In vitro conservation of plant biodiversity: approach in high school Abstract: Plants constitute the base of ecosystems and the need to conserve biodiversity is indispensable for society. The aim of this project was to raise awareness among students of a public high school in Rio de Janeiro, creating opportunity for undergraduate and graduate students to transmit concepts of biotechnology applied to plants’ conservation. The methodology consisted of lectures, videos documentaries and a technical visit to laboratories of the Plant Biotechnology Center of the Rio de Janeiro State University where students saw in vitro production and conservation of plants. The evaluation was based on the attendance and participation, as well as the comparison of the answers to questionnaires applied in the beginning and the end of the project. A large variation in the results between the classes was detected both in the frequency (41-68%) and in the percentage of correct answers in the initial (42-57%) and final (53-68%) questionnaires. However, all evaluated classes showed higher performance after participating in the project, indicating a positive effect on specific issues of biotechnology and its application in biodiversity conservation. Keywords: Sustainable Development, In Vitro Culture, Biology Teaching, Plant Biotechnology     Conservación in vitro de biodiversidad vegetal: enfoque en la enseñanza media Resumen: Las plantas constituyen la base de los ecosistemas y la necesidad de conservar la biodiversidad es imprescindible para la sociedad. El objetivo de este proyecto fue concientizar a los alumnos de la enseñanza media pública de esta realidad, generando oportunidad para graduandos y post-graduandos transmitir conceptos de biotecnología aplicada a la conservación de plantas. La metodología consistió de conferencias, vídeos y visita técnica a los laboratorios del Núcleo de Biotecnología Vegetal de la Universidad del Estado de Río de Janeiro, donde los alumnos acompañaron la producción y la conservación de plantas in vitro. La evaluación se basó en la comparación de las respuestas a un cuestionario aplicado al principio y al final del proyecto. Se detectó una gran variación en los resultados entre las clases, tanto en la frecuencia (41-68%), como en el porcentaje de aciertos en los cuestionarios inicial (42-57%) y final (53-68%). Sin embargo, todas las clases evaluadas presentaron mayor rendimiento después de la aplicación en el proyecto, indicando un efecto positivo en las cuestiones específicas de Biotecnología y su aplicación en la conservación de la Biodiversidad. Palabras-clave: Desarrollo Sostenible, Cultivo In Vitro, Enseñanza de Biología, Biotecnología Vegeta

Rescue of a non-viable accession and RAPD analysis of recovered plants of Arachis retusa

A regeneração in vitro de Arachis retusa foi avaliada visando à renovação e conservação de germoplasma. A estabilidade genética de plantas derivadas de eixos embrionários e segmentos apicais foi avaliada por RAPD. Foram analisados dez oligonucleotídeos decâmeros arbitrários, dos quais cinco foram selecionados. Noventa regiões genômicas foram avaliadas, com uma média de 18 loci por clone. Todos os segmentos amplificados foram monomórficos. Estes resultados indicam que as plantas são geneticamente estáveis nas regiões genômicas examinadas e que ambos os processos são adequados para a conservação in vitro do germoplasma de Arachis.In vitro regeneration of Arachis retusa was examined for the purpose of germplasm renewal and conservation. Random amplified polymorphic DNA (RAPD) fingerprinting was used to evaluate the genetic stability of plants derived from embryo axes and apical segments. Ten arbitrary decamer primers were screened and five of them were selected. Ninety genomic regions were evaluated, with an average of 18 loci per clone. All amplified segments were monomorphic. The results indicate that recovered plants are genetically stable at the assessed genomic regions and that both regeneration processes are suitable for in vitro germplasm preservation of Arachis species