Estudos sobre indução, manutenção e estabilidade genética da calogênese in vitro de Caesalpinia echinata Lam. (Pau-brasil)

Abstract

The Caesalpinia echinata Lam (Brazil wood) was the first protagonist of exploitation cycles in the history of Brazil, which contributed to the reduction of the Atlantic forest. His state of endangered species (IUCN, 2008; BRASIL, 2008) is due not only to the intense physical extraction and reduction of its range, but the persistent gaps in terms of knowledge about their spread (Barbedo et al., 2002). The plant tissue culture is an important technique and has been used successfully in the propagation of several species, including woody. Brunetta, in 2006, notes the same situation as found by Lee and Rao in 1988 that despite the success of in vitro propagation of woody plants in temperate climates, few studies employing this technique in tropical tree species. Callus formation is a basic step for the development of systems for mass propagation of plants through organogenesis or somatic embryogenesis, and is also useful in the production of cells for genetic manipulations, such as somatic hybridization, genetic transformation and poliploidizations. Besides these aspects, the callus culture may be used to obtain cell suspension, to study cell development and synthesis of products from primary and secondary metabolism. The combination effects of different concentrations and sources of auxin (2,4-D or NAA) and cytokinin (BAP or KIN) in modified MS medium, were evaluated in the induction of callus formation in different types of explants collected from different ontogenetic stages of Caesalpinia echinata Lam. The results demonstrate that the use of explants in the early stages significantly reduced infection rates and increased rates of callus formation. Callus formation was observed from the seed coat collected prior to dehiscence of the fruit, which probably originated from residual cells of the endosperm, or stomatal existing in this tissue. It was possible to induce callus and maintain viable for a long period, from explants contained the genotype of the stock plant, the genome resulting from fertilization of the egg cell and, possibly, the balance genomic 2m: 1p, a result of double fertilization which gives rise to the endosperm. The calluses were evaluated by AFLP (length polymorphism of amplified fragments), as to the possible occurrence of somaclonal variation in function: the source of explants, number of subcultures and maintenance time in vitro. The polyacrylamide gel electrophoresis of the fragments stained with silver nitrate produced few bands of good resolution and all monomorphic, showing that it is feasible using the AFLP technique in cell culture of the species, but should be tested in new combinations of nucleotides to amplify the power of the technique for the detection of somaclonal variation.