Method for manufacturing multiple amplification internal mark for four-bacteria PCR test
A technology of multiple amplification and amplification of internal standards, applied in biochemical equipment and methods, DNA preparation, recombinant DNA technology, etc., can solve problems such as unavailability, and achieve the effect of improving accuracy
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[0030] 1. Design of specific detection primers
[0031] The known specific genes of Staphylococcus aureus, Salmonella, Listeria monocytogenes and Vibrio parahaemolyticus were analyzed by bioinformatics, and the target genes vicK, invA, hlyA and toxR were selected for detection. The sequences of vicK, invA, hlyA, and toxR genes were respectively compared with other microorganisms through the public BLAST software in Genbank (an existing technology, shared free of charge), and sequence segments with higher specificity were selected. Then use the software Primer 5.0 (commercially available, Premier, Canada) to design a pair of internal standard primers in this specific sequence. The primer sequences are as follows:
[0032] 1. Primers for detection of Staphylococcus aureus (vicKF / vicKR)
[0033] vicKF: 5'- CGCAGGCTAATACTGAAAG -3'
[0034] vic KR: 5'- TTCTGTTTCTTCACGGGTA -3'
[0035] (a) Detect the sequence characteristics of the target gene:
[0036] * Length: 512 bp
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