关键词: G-四联体；聚合酶链式反应扩增；沙门氏菌；可视化检测

Abstract: In order to establish a visual detection method for Salmonella using G-guadruplex-generating polymerase chain reaction (PCR), the upstream and downstream primers containing G-quadruplex complementary sequences at the 5’ end were designed according to the Salmonella-specific gene invH, which served as the target gene. Double-stranded DNA contained a large number of G-quadruplexes was obtained after specific amplification of the target sequences by PCR, and bound to hemin after being denaturated to generate DNAzyme with peroxidase-like activity. DNAzyme was able to catalyze the reaction between H2O2 and 2,2’-azino-bis (3-ethylbenzothiazoline-6-sulfonic acid) diammonium salt, resulting in color change from colorless to green and thus allowing visual detection of Salmonella. Under the optimized detection conditions, there was a good linear relationship between the logarithm of Salmonella genome concentration (y) and absorbance at 421 nm (x), which was fitted to the equation: y = 0.129 9x + 0.217 9 (R2 = 0.994 5) with a linear range of 0.07–771.6 ng/μL, and the sensitivity was 0.07 ng/μL. The specificity analysis showed that this method was suitable for the detection of Salmonella, and could be successfully applied to artificially contaminated milk samples, with a detection limit of 1.2 × 102 CFU/mL. By using this method, 30 commercial samples were tested and the results were consistent with those obtained by the national standard detection method. This study provides a new strategy for the detection of foodborne pathogens.

Key words: G-quadruplex; polymerase chain reaction; Salmonella enteriditis; visual detection