Marine vibrio multiple PCR reaction reagent kit and detecting method thereof

Abstract

The present invention discloses one kind of multiple PCR reaction reagent kit for detecting marine vibrio and its detection method. The multiple PCR reaction reagent kit includes PCR reaction solution compounded with 10xPCR reaction buffering solution containing Mg2+, dNTP, Hot star Taq enzyme, pathogenetic vibrio DNA primer and sterilized ion solution. The pathogenetic vibrio DNA primer includes DNA primers of three kinds of vibrios. The present invention may be used in detecting these three kinds of vibrios in short time and lowered cost and is favorable to the diagnosis of vibriosis of marine animal.

Description

Technical field [0001] The invention relates to a detection technology for target DNA fragments, in particular to a multiple PCR reaction kit of marine vibrio and a detection method thereof. Background technique [0002] Vibriosis caused by bacteria of the genus Vibrio has severely harmed the marine aquaculture industry. Among marine Vibrio species, Vibrio alginolyticus, Vibrio harveyi and Vibrio parahaemolyticus are the main pathogens, which can infect grouper, red sea bream, perch, large yellow croaker, Japanese flounder and turbot. It can also infect crustaceans such as shrimp, portunus trituberculatus, scylla serrata, as well as shellfish such as clams, abalones, and oysters. Vibriosis has the characteristics of rapid onset, high infectivity and high mortality. Marine animals with vibriosis can develop epidermal ulcers, tail rot, ascites, and eye rot. Therefore, vibriosis causes great harm to the breeding marine animals and causes great economic losses in the marine aquacultu...

AI Technical Summary

Problems solved by technology

Vibrio alginolyticus, Vibrio harveyi and Vibrio parahaemolyticus among the marine vibrio are the main pathogenic bacteria, which can infect fish such as grouper, red sea bream, perch, large yellow croaker, flounder and turbot It can also infect crustaceans such as shrimp, Portunus trituberculatus, and Scylla serrata, as well as shellfish such as clams, abalone, and oysters. Vibriosis has the characteristics of rapid onset, high infectivity, and high mortality. Infection Marine animals with vibriosis will have symptoms such as skin ulcers, rotten tails, ascites, and rotten eyes. Therefore, vibriosis causes great harm to marine animals and leads to great economic losses in the mariculture industry.

[0003] Accurate and rapid detection of the pathogenic bacteria of vibriosis is the basis for drug prevention and control. Using polymerase chain reaction (PCR reaction) to detect the pathogenic bacteria of vibriosis has the advantages of rapidity, sensitivity and strong specificity. PCR The reaction kit is equipped with 10×PCR reaction buffer (containing Mg 2+ ), dNTP, Hot star Taq enzyme, DNA primers of pathogenic bacteria and sterilized deionized water mixed reaction solution, the detection method is to extract the DNA of the pathogenic bacteria of vibriosis, and add the DNA to the PCR reaction The kit performs an amplification reaction, and then electrophoresis is performed on the amplified product. By comparing the amplified product with the electrophoresis results of the positive standard and negative control, it is judged whether there is a pathogenic strain of the DNA primer, so as to find out the vibriosis pathogenic bacteria species; but generally only contain a single pair of primers of a pathogenic bacteria species in the existing PCR reaction kit, so each PCR reaction kit can only amplify the DNA of a pathogenic bacteria species, i.e. Only one pathogenic strain can be detected at a time, and the pathogenic strains that cause vibriosis are various, and the symptoms of vibriosis are also diverse, so that the DNA of which pathogenic strain is used can be selected. The detection of the primer kit has brought difficulties, and often requires multiple tests to find the pathogenic bacteria. Some vibriosis is co-infected by several marine Vibrio, so it is necessary to use different reagents for several pathogenic bacteria. PCR reaction kits are used to detect several pathogenic strains of vibriosis infection, which prolongs the detection of pathogenic strains, increases detection costs, and delays the prevention and control time of vibriosis, resulting in The spread of vibriosis infection, which leads to the aggravation of the condition of marine animals infected with vibriosis and more marine animals infected with vibriosis, causing significant economic losses in the mariculture industry

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