Universal linker Polymerase Chain Reaction-triggered Strand Displacement Amplification visual biosensor for ultra-sensitive detection of Salmonella

Abstract Salmonella is a principal causal agent of pathogenic outbreaks via food. A universal, highly sensitive and visual Salmonella detection method was proposed in this paper, based on a universal linker PCR (UL-PCR)-triggered Strand Displacement Amplification (SDA). In this research, the UL-PCR achieved the primary amplification. The universal linker primer was ingeniously designed and composed of two parts, one of which was the binding sequence of the target, and the other was the universal linker. Complementary sequences of the G-quadruplex and the nicking endonuclease recognition sequence were included in the universal linker. Therefore, the G-quadruplexes and nicking sites were successfully introduced into the UL-PCR products, providing a basis for further SDA triggering. SDA achieved the secondary signal amplification and generated a large amount of label-free DNAzymes. Following SDA, DNAzymes catalyzed 3,3’,5,5’-tetramethylaniline (TMB) into colored compounds visible to the naked eye. We obtained the best experimental conditions by univariate analysis. Under optimal conditions, this proposed universal label-free method could detect Salmonella genome at level as low as 22 copies mL-1, with an excellent linear range between 102 copies mL-1 and 107 copies mL-1. And the limit of quantification (LOQ) was 102 copies mL-1. This strategy shows promise for broad applications.