Estandarización de la técnica de electroforesis en gel con gradiente desnaturalizante (DGGE) para el análisis de poblaciones microbianas en bioreactores de cultivo mixto

Abstract

This work was the standardization of DGGE from different bioreactors with mixed cultures of micro-organisms, for which a method of DNA extraction for the additional amplification of 16S rRNA gene of the primers 338F and 518R generated an Amplicon of the region V3 of the gene by PCR, a method that was also standardized at the laboratory for further analysis by the technique are normalized DGGE. Is found that the temperature of annealed is a part fundamental for the development of the PCR since is the time in which is generates it hybridization between them primers and the chain of DNA, the Protocol TDown is a technical important, but reduces significantly the selectivity of them primers in the time of the amplification of the gene of interest for samples of mixed cultures obtained DMSO proved to be the best reagent for the optimization of the PCR, electrophoresis in polyacrylamide gel, ethidium bromide is a but you cannot add previously in the gel, if it is not that the subsequent staining is required with the solution, the PCR is a sensitive technique to several factors , but is very useful for the characterization of microorganisms due to its high selectivity. In terms of the DGGE, for complete standardization of the method, it is necessary to do more repetitions with different samples to validate it fully.