Double PCR primer, double PCR detection method and double PCR detection kit for grouper iridovirus

Abstract

The invention provides a double PCR primer, a double PCR detection method and a double PCR detection kit for grouper iridovirus. For the kit provided by the invention, the double PCR primer is designed according to major capsid protein (MCP) gene sequences of the genus megalocytivirus of grouper iridovirus (GIV-M) and the genus ranavirus of grouper iridovirus (GIV-R), a method for synchronously detecting GIV-M and GIV-R through a single tube is established, an amplification system and a method are optimized, and the high-specificity, high-sensitivity and rapid synchronous detection for GIV-M and GIV-R is finally realized.

Description

technical field [0001] The invention relates to the technical field of PCR, in particular to a double PCR primer, a detection method and a kit for grouper iridescent virus. Background technique [0002] Grouper is a rare marine fish with global distribution and has important economic value. China is the main producer of grouper in the world. Since 2014, the total annual output has been stable at about 100,000 tons (2016 China Fishery Statistical Yearbook), accounting for about 60% of the world's total grouper production, with an output value of more than 10 billion Yuan Renminbi. However, with the rapid development of grouper aquaculture, diseases occur more and more frequently, among which, viral diseases are particularly serious, which has become one of the bottlenecks restricting the healthy development of grouper industry. [0003] Grouper iridovirus (GIV) is a serious pathogen in grouper farming. It can infect more than 30 species of grouper and cause serious death. I...

AI Technical Summary

Problems solved by technology

Although these methods can detect fish viruses, they all have pathogenic defects: the detection sensitivity of nucleic acid probe method is low, less than 10 5 The copied virus cannot be detected, which cannot meet the needs of disease prevention and control; routine PCR, RT-PCR, fluorescent quantitative PCR, loop-mediated isothermal amplification (LAMP) and other methods, due to mutual interference of primers, the amplification of each viral gene must be It is carried out in different reaction tubes, so usually only one or several viruses can be detected, and the above-mentioned multiple fish viruses cannot be detected simultaneously and quickly. The detection efficiency is low, and it cannot meet the multi-virus screening requirements of unknown pathogenic samples. ; Although the nylon membrane gene chip method can simultaneously detect 17 kinds of fish pathogens, it still relies on conventional PCR and RT-PCR methods, and needs to amplify the pathogens separately in multiple reaction tubes, rather than in the same reaction tube. Each pathogen is amplified, so the workload is heavy, and operation errors are prone to occur; and this method uses nylon membrane dot hybridization technology to detect the amplification results, and uses naked eye observation to judge the detection results. The sensitivity is low and it is not easy to standardize, and the practical application range is limited.

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