Kit capable of rapidly detecting staphylococcus aureus in meat and application
A staphylococcus and golden yellow technology, applied in the field of food safety, can solve the problems of difficult inhibition and low requirement of DNA template purity, achieve good non-specific amplification and ensure specificity
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[0032] Example 1
[0033] 1. Experimental method
[0034] 1.1 The cultivation of different strains and the extraction of DNA templates
[0035] Take Staphylococcus aureus ATCC13565 as the standard strain, 3 strains of Staphylococcus aureus (ATCC25923, CMCC26074, CMCC26075) as the reference strain, and other 26 non-Staphylococcus aureus strains for specific experiments. Staphylococcus aureus and other strains were activated and cultured in NB and compound enrichment medium BPW, and cultured overnight at 37°C. Separately draw 1 mL of the bacterial suspension in the logarithmic growth phase, extract bacterial DNA with a bacterial genome extraction kit, and store the obtained DNA template at -20°C for use.
[0036] 1.2 Design and synthesis of LAMP primers
[0037] The Staphylococcus aureus heat-resistant nuclease gene (nuc) was found in GenBank. This gene has good specificity and is highly conservative. Therefore, this article uses this specific gene nuc sequence for primer design. Usin...
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[0070] Example 2:
[0071] Collect 10 pieces of fresh pork, beef, mutton, chicken and their meat products, and use the following kits and detection methods for detection.
[0072] The kit of the present invention is composed of 25 parts of LAMP amplification reaction solution, 1 part of negative control solution and 1 part of positive control according to the number of parts by volume. The LAMP amplification reaction system is composed of a LAMP primer set, Bst 2.0 WarmStart DNA polymerase and an amplification reaction solution. In the LAMP primer set, the concentration ratio of the outer primer to the inner primer is 1:6. Among them, each 25μL LAMP amplification reaction system consists of 0.4μM F3 and B3, 2.4μM FIP and BIP, 0.9μL 8U Bst 2.0 WarmStart DNA polymerase, 4mM MgCl2, 2mM dNTPs, 0.4M betaine, 2.5μL 10×Buffer and It consists of 2μL of template DNA and the rest is made up of sterilized double distilled water. The positive control solution is a genomic DNA solution extra...
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