FOOD SCIENCE ›› 2022, Vol. 43 ›› Issue (12): 325-328.doi: 10.7506/spkx1002-6630-20210304-060

• Safety Detection • Previous Articles    

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

LIU Jianhui, ZHANG Xianzhou, ZHANG Yunzhe, LI Lanru, WANG Hongjing, GAO Jie, GENG Fengzhen, TAN Jianxin   

  1. (1. College of Food Science and Technology, Hebei Agricultural University, Baoding 071000, China;2. Hebei Tourism College, Chengde 067000, China; 3. Affiliated Hospital of Hebei University, Baoding 071000, China)
  • Published:2022-07-01

Abstract: 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 + 2.872 3 (R2 = 0.992 5) 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.

Key words: G-quadruplet; polymerase chain reaction; rolling circle amplification; Salmonella; fluorescence signal detection

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