Morfogênese in vitro a partir de segmentos de hipocótilos e de raízes de urucum (Bixa orellana L.)

• Autor(a) principal: Matos, Elyabe Monteiro de
• Data de Publicação: 2009
• Tipo de documento: Dissertação
• Idioma: por
• Título da fonte: LOCUS Repositório Institucional da UFV
• Texto Completo: http://locus.ufv.br/handle/123456789/2505
• topic: Morfogênese in vitro; Bixa orellana; Hipocótilos; Raízes; Morphogenesis in vitro; Bixa orellana; Hypocotyl; Roots; CNPQ::CIENCIAS BIOLOGICAS::BOTANICA

Bixa orellana (Bixaceae) popularly known as annatto is a wood species native from Tropical America, and widely distributed in all Brazilian territory. The seeds are a wealthy source of the carotenoids bixin and norbixin, largely used in food, cosmetic, and pharmaceutical industries, and presenting high economic interest. The development of in vitro regeneration protocols is important as a mean to generate new alternatives for obtainment of genetically modified plants with genes of agronomic interest and those associated to the biosynthesis of carotenoids, allowing a better understanding and manipulation their biosynthetic pathways. The goals of this work had been to evaluate the influence of zeatin pulses, to test different medium of culture in organogenesis of annatto, beyond testing different concentrations of Pluronic® F-68 and to analyze the combinations of mannose: sucrose in morphogenesis of annatto. In the present work, different exposure periods to growth regulators using hypocotyl segments were assayed. In average, 8.8 shoots were obtained per explant and high organogenesis frequency (80 %), when explants were cultured on induction medium (MS supplemented with 4.56 µM zeatin) during 6 days, and transferred to MS medium devoid of growth regulators. However, adventitious regeneration frequencies around 60 % were achieved when explants remained for 2 or 3 days on induction medium. Regarding media formulation used, MS and JADS propitiated the highest organogenic frequencies for genotype M2 (98 and 90 %, respectively), whereas WPM led to the lowest regeneration frequencies (6 %). DKW medium enabled intermediate regeneration frequencies ranging from 70 to 80 %. For Piave Vermelha it was observed that JADS medium provided the highest frequencies, as compared to other media (68 %). It was noticed genotype-dependent regeneration responses. The non-ionic surfactant Pluronic® F-68 was tested in different combinations with zeatin 4.56 µM, for both hypocotyl and root segment explants. No significant variations were observed for Pluronic® F-68 used for M2 , but the lowest frequencies were obtained at 0.001 % (82 %). However, for Piave Vermelha the lower surfactant concentrations (0.001 and 0.1 %) led to greater shooting frequencies in hypocotyl segments (28 and 12 %, respectively). For M2 root explants, Pluronic® F-68 at 0.001 e 0.5 %, promoted the highest adventitious shoot numbers (43 e 40 %, respectively); root explants of Piave Vermelha did not display any response to the same surfactant treatments. The use of an alternative carbon source (mannose) as a selective agent in annatto genetic transformation by means of pmi (phosphomannose isomerase) gene was evaluated for both hypocotyl and root explants. Annatto explants were not able to metabolize the mannose present alone or in combination with sucrose, on zeatin- supplemented (4.56 µM) JADS medium. This way, no regenerative response was achieved when mannose was the sole carbon source; sucrose, even at the lowest concentration (21.9 mM) promoted shoot morphogenesis. Root explants from Piave Vermelha did not show any response to the different carbon source treatments, however M2 root explants showed the higher number of shoots (7.2) when 75 % of the mannose were replaced by sucrose. Therefore, analyzing the overall behavior of both genotypes it can be concluded that interespecific variation, the mineral composition of the medium, the supplementation with organic/inorganic substances, growth regulators, gelling agents, and the type of explants must be taken into account when targeting the in vitro morphogenesis of annatto, aiming at the optimization of in vitro responses of this species.