Influence of buffer systems on the interaction between a random DNA library and Salmonella typhimurium cells.
DOI:
https://doi.org/10.15649/2346075X.3556Keywords:
Random DNA library, Cell-SELEX, S. typhimurium, MgCl₂Abstract
Introduction. Salmonella enterica subsp. enterica ser. Typhimurium (S. typhimurium) is a gram negative bacterium of significant public health concern due to its ability to be transmitted through food and is the cause of several diseases collectively known as salmonellosis. The development of rapid and specific detection systems for this microorganism is crucial to strengthen food quality control programs and the prevention of infectious outbreaks. This need underscores the importance of studying novel diagnostic approaches, such as those based on aptamers, which are typically generated by SELEX technique. The parameters for this technique are defined by individual research groups and directly affect recognition capabilities of the aptamers for their targets. Objective. In this study, we investigated the influence of three different buffer systems and varying concentrations of magnesium chloride (MgCl₂) on the binding capacity of a single-stranded random DNA library to S. typhimurium cells. Materials and Methods. S. typhimurium cells were incubated with random DNA sequences in the presence of phosphate-buffered saline (PBS), Tris-buffered saline (TBS), or Tris-KNO3 buffers with varying concentrations of magnesium chloride (MgCl₂). Unbound or weakly bound sequences were separated by centrifugation and quantified by qPCR. Statistical differences between experimental conditions were analyzed using analysis of variance (ANOVA). Results. The selection of buffer system did not significantly affect the recovery of DNA sequences complexed to S. typhimurium cells at a constant MgCl₂ concentration. However, changes in MgCl₂ concentration markedly influenced the amount of sequences bound to S. typhimurium cells. Conclusions. The interaction between DNA sequences and S. typhimurium cells was maximized at a MgCl₂ concentration of 2.5 mM in TBS buffer.
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