Estudo epigenético e molecular aplicados à biotecnologia vegetal

Abstract

The present study investigated two distinct approaches within plant biotechnology: the epigenetic analysis of two cultivars of Vigna unguiculata [L.] Walp. infected with Cowpea Severe Mosaic Virus (CPSMV), and the interaction between the R-phycoerythrin (R-PE) protein from the red macroalga Solieria filiformis and DNA as a potential biotechnological product. In the first chapter, the objective was to identify DNA methylation patterns in contrasting cowpea cultivars infected with CPSMV using the MSAP technique. The results showed variations in methylation profiles over time following infection, with a higher abundance of fragments observed 48 hours after inoculation. The methylation levels in the 144h group showed a general decrease in methylation in profiles II and III when compared to those at 48h. The predominance of demethylation in the resistant cultivar may be associated with the activation of defense-related genes, whereas the predominance of increased methylation in the susceptible cultivar may be linked to redox imbalance and a lower survival rate. However, to precisely determine how methylation influences plant resistance or susceptibility, further studies with complementary analyses, such as sequencing, are required. The second chapter investigated the ability of R-PE to interact with DNA molecules of different conformations through agarose and polyacrylamide gel electrophoresis. The experiments revealed that native R-PE maintained its fluorescence, while the denatured form showed reduced intensity. The presence of high-molecular-weight bands in samples containing R-PE and DNA suggests slight protein degradation or a possible protein-DNA interaction, indicating potential application as an intercalating agent. Additionally, bands corresponding to the pENTR GUS plasmid were observed on the agarose gel through fluorescence emission from R-phycoerythrin interacting with the DNA. Based on the observed indications of interaction, it is necessary to repeat the gels using negative controls for both DNA and protein in order to rule out false positives and confirm the interaction.