Estabelecimento, multiplicação, calogênese, organogênese in vitro e análise da diversidade genética em acessos de Eugenia involucrata DC.

Abstract

The cerejeira-do-Rio-Grande (Eugenia involucrata DC.) is a Brazilian forest species with traits of economic interest, especially in the forestry, horticulture, landscape, environmental and medicinal sectors. However, it has recalcitrant seeds, which begin to lose viability after two weeks of gathering. Moreover, doesn t known how its genetic variability is distributed. This study aimed to developed methodologies for in vitro culture of E. involucrata and evaluate the genetic variability in accessions maintained in situ and ex situ. In vitro propagation, besides desinfestation, we evaluated aspects of in vitro growth and development after the inoculation of the apical and nodal segments in different nutritive media. We also analyzed the effect of introduction of different growth regulators in the optimization of in vitro multiplication. For callus induction in vitro were evaluated methods of superficial desinfestation, ways of introduction of plant growth regulators, the effect of different light regimes, of the use of antioxidants and of the different cytokinins. The formation of callus on different aspects were evaluated. We tested different methods for extraction of genomic DNA from this specie using different plant tissues. We performed analysis of acess coming from populations maintained in situ and ex situ by the use of markers. It was possible to establish aseptic cultures of E. involucrate. The establishment in vitro can be accomplished with the apical and nodal segments, however, nodal segments offer the best results. The ½ MS is the most appropriate nutritive medium and helps in rooting. The use of high concentrations of TDZ associated with ANA enables shoot multiplication and the emergence of shoots from nodal segments. GA3 does not promote the elongation of shoots and is toxic in high concentrations. Callus formation was obtained in leaf discs of E. involucrata. The best way to grow is maintaining the explants in abaxial position and in the dark. Combinations of auxins and cytokinins generate the highest percentage of callus formation. The association of 2,4-D and TDZ promotes better callus formation and callus candidates to embyogeneis. It is possible to extract good quality genomic DNA from E. involucrata leaves and cambia, using the maceration with the extraction buffer concentrate to 10%. Analysis with molecular markers was efficient in the stratification of genetic variability in accessions of cherry. It was observed that the variability is evenly distributed between the accessions in different in situ populations and that in the ex situ population assessed the variability is not satisfactory. It was possible to generate consistent information to assist in the cultivation and in planning of breeding programs of the species.