Multiplex PCR diagnosis kit for three pathogenic bacteria in milk

Abstract

The invention discloses a multiplex PCR diagnosis kit for three pathogenic bacteria in milk. Primers of the kit are specific primers in a staphylococcus aureus femB gene sequence, an escherichia coliphoA gene sequence and a salmonella invA gene sequence. Through optimization of multiplex PCR reaction and an optimization experiment, the optimal amplification procedure for multiplex PCR of the kitis determined to be 5 min at 95 DEG C, 30 s at 95 DEG C, 35 s at 50-60 DEG C, 45 s at 72 DEG C, amplification for 30 cycles, and final extension at 72 DEG C for 10 min. The multiplex kit disclosed bythe invention can rapidly and accurately detect and identify separate or mixed infection of staphylococcus aureus, salmonella and escherichia coli in a clinical milk sample. The experimental result proves that the kit is short in detection time, good in sensitivity and high in specificity and has a broad application prospect.

Description

technical field [0001] The invention belongs to the technical field of detection of food-borne pathogenic bacteria, in particular to a multiple PCR diagnostic kit for three kinds of pathogenic bacteria in milk. Background technique [0002] Foodborne pathogens are one of the main factors that cause human health and food safety, and microbial contamination is an important factor that leads to milk quality problems, so pathogenic bacteria in milk have become the focus of consumers' attention. However, my country's quality requirements for raw milk are far lower than those of developed dairy countries. The new national standard for dairy products in my country only stipulates the minimum total number of bacteria detected in raw milk (less than 2×10 6 individual / mL), there is no regulation on the specific bacterial species in milk, which ignores the detection of pathogenic bacteria. Escherichia coli, Staphylococcus aureus, and Salmonella are the main pathogenic bacteria derive...

AI Technical Summary

Problems solved by technology

Although this patented technology realizes the simultaneous detection of Staphylococcus aureus, Escherichia coli, and Salmonella, the fluorescent quantitative PCR consumables are expensive, and it has not been popularized in county-level grassroots and farms at present, and the operating technology requires high requirements for the cleanliness of the operating environment. High, prone to false positives due to environmental pollution

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