Selection, identification and application of DNA aptamers for the detection of Bifidobacterium breve

In the present study, a single-stranded DNA (ssDNA) aptamer binding to Bifidobacterium breve with high avidity and selectivity was selected through a whole-bacterium-based systemic evolution of ligands using an exponential enrichment (SELEX) process. Following 12 rounds of selection specific for B. breve, three FAM-labeled aptamers were chosen for flow cytometry analysis and the results revealed that all three aptamers possessed a high binding affinity for B. breve. To obtain the optimal sequence, sequence truncation experiments of these three aptamers were conducted. An aptamer variant BB16-11f with high affinity and selectivity was acquired. In addition, the dissociation constant was significantly reduced to 18.66 1.41 nM. Furthermore, an enzyme linked aptamer assay was developed to prove the potential application of the aptamer BB16-11f in the detection of B. breve. The results showed that the colorimetric assay had a linear relationship between the absorbance at 450 nm and the concentrations of B. breve ranging from 10 cfu mL 1 to 10 cfu mL 1 with a correlation coefficient of 0.98. The limit of detection (LOD) of the assay was 1000 cfu mL . Additionally, the developed method was also successfully used to detect B. breve in a milk environment. Taken together, we hold that the developed colorimetric bioassay based on the aptamer BB16-11f is a promising method for the detection of B. breve.