RESUMO
Gamma radiation (60Co)-induced mutagenesis offers an alternative to develop rice lines by accelerating the spontaneous mutation process and increasing the pool of allelic variants available for breeding. Ionizing radiation works by direct or indirect damage to DNA and subsequent mutations. The technique can take advantage of in vitro protocols to optimize resources and accelerate the development of traits. This is achieved by exposing mutants to a selection agent of interest in controlled conditions and evaluating large numbers of plants in reduced areas. This chapter describes the protocol for establishing gamma radiation dosimetry and in vitro protocols for optimization at the laboratory level using seeds as the starting material, followed by embryogenic cell cultures, somatic embryogenesis, and regeneration. The final product of the protocol is a genetically homogeneous population of Oryza sativa that can be evaluated for breeding against abiotic and biotic stresses.
Assuntos
Raios gama , Mutagênese , Oryza , Sementes , Oryza/genética , Oryza/efeitos da radiação , Oryza/crescimento & desenvolvimento , Mutagênese/efeitos da radiação , Sementes/genética , Sementes/efeitos da radiação , Sementes/crescimento & desenvolvimento , Regeneração/genética , Técnicas de Embriogênese Somática de Plantas/métodosRESUMO
The development of gamma ray-mutated rice lines is a solution for introducing genetic variability in indica rice varieties already being used by farmers. In vitro gamma ray (60Co) mutagenesis reduces chimeras and allows for a faster selection of desirable traits but requires the optimization of the laboratory procedure. The objectives of the present work were sequencing of matK and rbcL, the in vitro establishment of recalcitrant rice embryogenic calli, the determination of their sensitivity to gamma radiation, and optimization of the generation procedure. All sequenced genes matched perfectly with previously reported matK and rbcL O. sativa genes. Embryogenic calli induction improved using MS medium containing 2 mg L-1 2,4-D, and regeneration was achieved with MS medium with 3 mg L-1 BA and 0.5 mg L-1 NAA. The optimized radiation condition was 60 Gy, (LD20 = 64 Gy) with 83% regeneration. An immersion system (RITA®, Saint-Mathieu-de-Tréviers, France) of either 60 or 120 s every 8 h allowed systematic and homogeneous total regeneration of the recalcitrant line. Other well-known recalcitrant cultivars, CR1821 and CR1113, also had improved regeneration in the immersion system. To our knowledge, this is the first study reporting the use of an immersion system to allow for the regeneration of gamma-ray mutants from recalcitrant indica rice materials.
RESUMO
Abstract The objective of this study was to identify the emetic metabolites in different parts of the P. ipecacuanha, a plant with emetic properties. Partial phytochemical analysis was performed to determine the presence of emetine and cephaeline in leaves, stems and roots. Both alkaloids were detected in the three plant parts analyzed. Highest alkaloid content was found in roots (8.55 mg/g), followed by stems (4.05 mg/g), and the lowest was found in leaves (2.4 mg/g). The cephaeline content (8.35 mg/g) was higher than that of emetine (6.65 mg/g) in all the three organs analyzed. Toxicity analysis of the crude extract showed a LD50 of 500 mg/kg.
Resumen El objetivo de este estudio fue identificar los metabolitos eméticos en diferentes partes de P.ipecacuanha, una planta con propiedades eméticas. Se realizó un análisis fitoquímico parcial, donde se determinó la presencia de emetina y cefalina en hojas, tallos y raíces. Ambos alcaloides se detectaron en las tres partes de la planta analizadas. El mayor contenido de alcaloides se encontró en las raíces (8.55 mg/g), seguido de los tallos (4.05 mg /g), y el más bajo se encontró en las hojas (2.4 mg/g). El contenido de cefalina (8,35 mg/g) fue mayor que el de emetina (6,65 mg/g) en los tres órganos analizados. El análisis de toxicidad del extracto crudo mostró una DL50 de 500 mg/kg.
Resumo O objetivo deste estudo foi identificar os metabólitos eméticos em diferentes partes do P. ipecacuanha, uma planta com propriedades eméticas. Foi realizada análise fitoquímica parcial para determinar a presença de emetina e cefelina nas folhas, caules e raízes. Ambos os alcalóides foram detectados nas três partes da planta analisadas. O maior teor de alcalóides foi encontrado nas raízes (8,55 mg/g), seguido dos caules (4,05 mg/g), e o menor foi encontrado nas folhas (2,4 mg/g). O conteúdo de cefhaelina (8,35 mg/g) foi superior ao de emetina (6,65 mg/g) nos três órgãos analisados. A análise de toxicidade do extrato bruto mostrou um LD50 de 500 mg/kg.
RESUMO
Experiencia práctica: socialización de conceptos, aplicaciones y beneficios de la biotecnología en Costa Rica. Los procesos biotecnológicos han acompañado a la humanidad desde el inicio de la civilización; prueba de esto es el uso de levaduras para preparar pan, vino y cerveza. La biotecnología también ha sido fundamental en el mejo ramiento de plantas y animales que son parte de la dieta humana; y las técnicas de cultivo in vitro han acelerado el proceso de obtener mejores cultivos para nuestra creciente población. El proyecto "Biotecnología para todos: socialización de conceptos, aplicaciones y beneficios" socializa los conceptos, aplicaciones y beneficios de la biotecnología entre educadores, grupos formadores de opinión, y productores de los sectores de agricultura y alimentos. El proyecto ha utilizado exitosamente nuevas tecnologías para lograr sus objetivos.
Biotechnological processes have accompanied humanity since the beginning of civilization; proof of this is the use of yeasts for the preparation of bread, wine and beer. Biotechnology has also been fundamental in the improvement of plants and animals that are part of the human diet; and in vitro culture techniques have accelerated the processes for obtaining better crops for our growing population. The project "Biotechnology for everyone: Socialization of concepts, applications and benefits" socializes the concepts, applications and benefits of biotechnology among educators, opinion-forming groups, and producers in the agricultural and food sectors. The project has successfully used new technologies to reach its goals.
RESUMO
This paper presents the development of cryopreservation protocols for zygotic embryos and apices of chayote (Sechium edule Jacq. Sw.), a tropical plant species with recalcitrant seeds. Zygotic embryos of two cultivars, Ccocro negro (CN) and Claudio (Cl) could withstand cryopreservation, with survival percentages of 10 and 30 %, after desiccation to 23 and 19 % moisture content (fresh weight basis), respectively. Apices sampled on in vitro plantlets of cultivars Cl, 13 and JM were successfully cryopreserved using a vitrification technique. Optimal conditions included the culture of mother-plants for 22 days on medium containing 0.3 M sucrose, culture of excised apices on the same medium for 1 day, loading of apices for 20 min with 2M glycerol + 0.4M glycerol, treatment with a series of diluted PVS2 solution (60 % PVS2 followed by 80 % PVS2 solution for 15 min (cultivar Cocoro Blanco [CB]) or 30 min (cultivars CN and Cl) at each concentration), rapid freezing and thawing, washing of shoot-tips with a 1.2 M sucrose solution, followed by recovery on media with progressively decreasing sucrose concentrations until the standard concentration of 0.1 M was reached. The highest survival percentages achieved ranged between 17 and 38 %, depending on the cultivar.
