Detection of Salmonella by G-quadruplex-Based PCR-RCA Double Amplification

In this study, a method to detect Salmonella was developed using polymerase chain reaction (PCR) combined with rolling circle amplification (RCA) with a dumbbell ring-shaped template into which a G-quadruplex complementary sequence was inserted. This method was based on PCR-RCA double amplification and the fact that thioflavin T (ThT) can specifically bind to the G-quadruplex, enhancing the fluorescence signal. Under the optimized detection conditions, the logarithmic concentration of Salmonella genomic DNA (x) had a good linear relationship with the fluorescence signal intensity at 486 nm (y). The regression equation was y = 2.101x + 2.8723 (R² = 0.9925) in a linear range of 17 fg/μL‒1.7 ng/μL. The specificity of the method was evaluated, revealing that it was suitable for the detection of Salmonella with a detection limit of 4.28 CFU/mL for artificially contaminated milk samples. This method, which has several advantages such as strong specificity, high sensitivity and low detection limit, provides a new method for the rapid detection of foodborne pathogens.