RESUMO

Jatropha curcas is an oilseed crop renowned for its tolerance to a diverse range of environmental stresses. In Brazil, this species is grown in semiarid regions where crop establishment requires a better understanding of the mechanisms underlying appropriate seed, seedling and plant behaviour under water restriction conditions. In this context, the objective of this study was to investigate the physiological and cytological profiles of J. curcas seeds in response to imbibition in water (control) and in polyethylene glycol solution (osmoticum). Seed germinability and reactivation of cell cycle events were assessed by means of different germination parameters and immunohistochemical detection of tubulin and microtubules, i.e. tubulin accumulation and microtubular cytoskeleton configurations in water imbibed seeds (control) and in seeds imbibed in the osmoticum. Immunohistochemical analysis revealed increasing accumulation of tubulin and appearance of microtubular cytoskeleton in seed embryo radicles imbibed in water from 48 h onwards. Mitotic microtubules were only visible in seeds imbibed in water, after radicle protrusion, as an indication of cell cycle reactivation and cell proliferation, with subsequent root development. Imbibition in osmoticum prevented accumulation of microtubules, i.e. activation of cell cycle, therefore germination could not be resumed. Osmoconditioned seeds were able to survive re-drying and could resume germination after re-imbibition in water, however, with lower germination performance, possibly due to acquisition of secondary dormancy. This study provides important insights into understanding of the physiological aspects of J. curcas seed germination in response to water restriction conditions.

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RESUMO

Actually, the germplasm of Jatropha spp. is conserved as whole plants in field collections. Under this storage method, the genetic resources are exposed to disease, pest and natural hazards such as human error, drought and weather damage. Besides, field genebanks are costly to maintain and with important requirements of trained personnel. Thus, the development of efficient techniques to ensure its safe conservation and regeneration is therefore of paramount importance. In this work we describe a method for Jatropha curcas seeds cryoexposure and seedling recovery after thawed. In a first experiment, an efficient protocol for in vitro plant recovery was carried out using zygotic embryo or seeds with or without coat. In a second experiment, desiccated seeds with or without coat were exposed to liquid nitrogen and evaluated after cryoexposure. Germination percentages were variable among treatments, and seeds demonstrated tolerance to liquid nitrogen exposure under certain conditions. Seeds of J. curcas presented up to 99.6% germination after seed coat removal. Seeds with coat cultured in vitro did not germinate, and were 60% contaminated. The germination of the zygotic embryos was significantly higher in the ½ MS medium (93.1%) than in WPM medium (76.2%), but from zygotic embryo, abnormal seedlings reached up to 99%. Seeds with coat exposed to liquid nitrogen showed 60% germination in culture after coat removal with good plant growth, and seeds cryopreserved without coat presented 82% germination, but seedlings showed a reduced vigor and a significant increase in abnormal plants. Seeds cultured in vitro with coat did not germinate, independently of cryoexposure or not. This study reports the first successful in vitro seedling recovery methodology for Jatropha curcas seeds, after a cryopreservation treatment, and is recommended as an efficient procedure for in vitro plant recovery, when seeds are conserved in germplasm banks by low or cryotemperatures.

Actualmente, el germoplasma de las especies de Jatropha ssp. se conserva como plantas enteras en las colecciones de campo. Bajo este método de almacenamiento, los recursos genéticos están expuestos a enfermedades, plagas y desastres naturales tales como el error humano, la sequía y las inclemencias del tiempo. Además, los bancos de germoplasma de campo son costosos de mantener y requieren bastante personal capacitado. Por lo tanto, el desarrollo de técnicas eficientes para asegurar su conservación segura así como su regeneración, es de suma importancia. En este trabajo se describe un método de recuperación para semillas y plántulas crioexpuestas de Jatropha curcas después de descongeladas. En un primer experimento, se llevó a cabo un protocolo eficiente para la recuperación de plantas in vitro mediante el uso de embriones cigóticos o semillas con o sin testa. En un segundo experimento, las semillas disecadas, con o sin testa fueron expuestas a nitrógeno líquido y se evaluaron después de la crioexposición. Los porcentajes de germinación fueron variables entre los tratamientos, y las semillas demostraron tolerancia a la exposición del nitrógeno líquido bajo ciertas condiciones. Las semillas de J. curcas presentaron hasta un 99.6% de germinación después de la eliminación de la testa. Las semillas con la testa cultivadas in vitro no germinaron, y el 60% se contaminaron. La germinación de los embriones cigóticos fue significativamente alta en el medio ½ MS (93.1%) en comparación con el medio WPM (76.2%), pero desde los embriones zigóticos, las plántulas anormales alcanzaron más del 99%. Semillas con la testa inmersa en nitrógeno líquido mostraron un 60% de germinacion en cultivos despúes de la remoción de la testa con un buen crecimiento de la planta, y las semillas criopreservadas sin testa presentaron un 82% de germinación, pero las plántulas mostraron un reducido vigor y un incremento significativo de plantas anormales. Semillas con testa cultivadas in vitro no germinaron, independientemente de la criopreservación o no. Este estudio reporta el primer éxito in vitro de una metodología de recuperación de plántulas para semillas de Jatropha curcas, después de un tratamiento de criopreservación, que se recomienda como un procedimiento eficaz para la recuperación de plantas in vitro, cuando las semillas se conservan en bancos de germoplasma a bajas o crio-temperaturas.

Assuntos