动物造模 z-bio 2021-06-22 377

　　Objective: To establish a loop-mediated isothermal amplification (LAMP) method for the detection of Salmonella faecalis tree shrews and to apply the method preliminarily.

　　Method: Design and synthesize specific LAMP primers based on the conservative sequence of Salmonella species-specific gene invA (infiltrating protein gene A). Set 10 reaction temperatures (57°C-66°C) and 10 reaction times (24-42 minutes) respectively for optimization to determine the specificity and sensitivity of the method. At the same time, a normal PCR experiment was performed to verify and compare this. Using the established LAMP method, 91 tree shrew loose stool samples were detected from wild sources.

　　Result: Under the optimized reaction conditions, the reaction temperature was 62°C and the reaction time was 34 minutes. The sensitivity of this method to Salmonella reaches 3.36×101 CFU/mL, which is 10-100 times that of conventional PCR methods. The gut microbiota of 10 tree shrews was tested by LAMP. Only Salmonella enteritidis and Salmonella B of the genus Salmonella were positive, and the rest were negative. 91 samples of wild tree shrew feces were tested for LAMP and the results were positive. 20.88? Can all reactions in the LAMP method be completed within 40 minutes? The result can be directly judged by visually observing the color change.

　　Conclusion: The established LAMP method is convenient, rapid, sensitive and specific, and can be used for large-scale rapid detection of Salmonella in tree shrew feces.