A multiple PCR primer used for simultaneously detecting five pathogenic bacteria and three virulence genes in marine, and a designing method thereof

Abstract

The invention relates to a multiple PCR primer used for simultaneously detecting five pathogenic bacteria and three virulence genes in marine, and a designing method thereof. The method includes: designing a multiple PCR primer composition capable of simultaneously detecting enterococcus faecalis, edwardsiella tarda, carnobacterium, bacillus thuringiensis, vibrio alginolyticus, an enterobacter cloacae gyrB gene, a vibrio parahaemolyticus tlh gene, and an enterococcus faecalis cylA gene by utilization of a primer designing software, screening primers dominant in competitive states, subjecting the primers dominant in the competitive states to amplification and verification by utilization of a PCR method, and finally removing primers weak in the competitive states in the amplification primers so as to obtain a multiple PCR primer composition. A detection method of the multiple PCR is also disclosed. Compared with the prior art, the multiple PCR primer, the designing method and the detection method are advantageous in that: the multiple PCR primer composition simultaneously detecting the five pathogenic bacteria and the three virulence genes in marine has advantages of simple and convenient operation, rapidness, strong specificity, high sensitivity, and the like.

Description

technical field [0001] The invention belongs to the field of microbial detection, and specifically relates to a method for simultaneously detecting marine Enterococcus faecalis, Edwardsiella tarda, Carnobacterium thuringiensis, Vibrio alginolyticus, Enterobacter cloacae gyrB gene, Vibrio parahaemolyticus tlh gene, fecal enterococci Multiplex PCR primers of coccus cylA gene and its design method. Background technique [0002] Enterococcus faecalis, Edwardsiella tarda, Carobacillus, Bacillus thuringiensis and Vibrio alginolyticus are pathogenic bacteria that can be transmitted through food and water. There are certain differences in their biological characteristics and pathogenicity, but they can all cause food poisoning and digestive system diseases. [0003] Enterococcus faecalis is ubiquitous in nature and can cause many diseases such as endocarditis, cholecystitis, meningitis, urinary tract infection and wound infection. It has low nutritional requirements and can also g...

AI Technical Summary

Problems solved by technology

This is because a single pair of primers can amplify a single template to amplify the target fragment with clear bands, but the phenomenon of band loss will occur after the amplification of various primers is mixed.

There is also a competitive relationship between the primers, and the strong primers may cover the weak primers, resulting in the loss of the target fragment

Even the interaction between the primers produces serious primer-dimers, resulting in the loss of the target fragment

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