Kit for rapidly detecting salmonella in meat and application thereof
A Salmonella and kit technology, applied in the field of kits for rapid detection of Salmonella in meat, can solve the problems of time-consuming and laborious instrument requirements, cumbersome detection of Salmonella, etc., and achieve the effects of high sensitivity, improved detection sensitivity, and strong specificity
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Embodiment 1
[0037] 1. Use bacterial genome kit to extract Salmonella DNA in meat samples to be tested
[0038] 2. Use the extract as a template for LAMP amplification, and the reaction system is 25 μL
[0039]
[0040]
[0041] LAMP amplification conditions are: react at 95°C for 5 minutes, quickly place at 4°C to add Bst enzyme, react at 65°C for 60 minutes, and inactivate the enzyme at 80°C for 2 minutes.
[0042] 3. Electrophoretic detection of amplified products
[0043] Take 5 μL of the amplified product for 1% agarose gel electrophoresis, electrophoresis at 110V for 40 min, and use the gel imaging system to observe the results ( figure 1 ).
[0044] This implementation method has simple requirements on instruments and equipment, can be completed only under water bath conditions, takes about 2 hours to complete, and has high detection sensitivity, with a detection limit of 9.8×10 1 CFU / mL.
Embodiment 2
[0046] 1. The DNA extraction of the meat sample to be tested is the same as in Example 1.
[0047] 2. LAMP was carried out using the extract as a template, and the reaction volume was 25 μL.
[0048]
[0049]
[0050] 3. Electrophoretic detection of amplified products
[0051] Take 5 μL of the amplified product for 1% agarose gel electrophoresis, electrophoresis at 110V for 40 min, and use the gel imaging system to observe the results.
[0052] This implementation method has simple requirements on instruments and equipment, only needs water bath conditions to complete, takes about 2 hours to complete, and has high detection sensitivity with a detection limit of 5.7×10 2 CFU / mL.
Embodiment 3
[0053] Embodiment 3 about the test of sensitivity
[0054] The method of the embodiment of the present invention 1 is carried out the test of sensitivity, specifically as follows:
[0055] Fresh pork was purchased from the local farmer's market, and it was confirmed by the national standard method before artificial contamination that it does not contain Salmonella. Take 25g pork sample and put it into a sterile homogeneous bag containing 225mL BPW, make a 1:10 dilution, artificially inoculate Salmonella, and incubate at 37°C for 12h. Adjust the cell concentration in the homogenate to add 10 7 , and then perform 10-fold serial dilution, so that the concentration of Salmonella is 10 7 CFU / mL~10 -1 CFU / mL, respectively extract the DNA of Salmonella in each group of artificially polluted meat pulp, the method is the same as in Example 1, and the extract is used as a template to carry out LAMP and PCR reactions, and the electrophoresis of the amplification result is as follows: ...
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