Method for fast detecting reverse transcription-polymerase chain reaction of paralichthys rhabdovirus

A technology of chain reaction and detection method, which is applied in the directions of microorganism-based methods, microbial determination/inspection, biochemical equipment and methods, etc. High sensitivity and accurate detection effect

Inactive Publication Date: 2010-01-13
孙颖杰 +4
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Problems solved by technology

Currently, fish virus detection methods mainly include three categories: cytological diagnostic technology, immunological diagnostic technology, and molecular biological diagnostic technology. , the detection cycle is long, and the sensitivity is low; immunological diagnostic techniques mainly include immunofluorescence detection and immune dot hybridization, which have the advantages of strong specificity and high sensitivity, but the method is quite cumbersome and is not suitable for detection of a large number of samples; Biological diagnostic techniques mainly include polymerase chain reaction (PCR), which is relatively fast and sensitive, and is currently widely used in the detection of fish viruses, such as infectious hematopoietic necrosis virus, koi herpes virus, infectious pancreatic necrosis virus, Red sea bream iridescent virus, etc. According to the search, there is no report on the detection of flounder rhabdovirus by RT-PCR technology

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  • Method for fast detecting reverse transcription-polymerase chain reaction of paralichthys rhabdovirus

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Embodiment 1

[0018] Example 1: The flounder rhabdovirus was detected by taking the flounder sample as an example.

[0019] The flounder sample was collected from a farm in the Yellow Coast. It had clinical symptoms of infection, including hyperemia or hemorrhage on the body surface and fins, distended abdomen, and ascites inside. Dissecting the fish body, the lamina propria of the muscle and intestinal mucosa are bleeding, and the connective tissue of the gonad is congested or bleeding.

[0020] The specific detection steps are as follows:

[0021] 1. Design specific primers:

[0022] In http: / / www.ncbi.nlm.nih.gov / Genbank / , search for the rhabdovirus sequence of the flounder, and use the existing DNA LASERGENE 7.1 software to analyze and compare the conservation of each gene sequence. The results show that the rhabdovirus of the flounder The glycoprotein gene sequence of the virus is relatively conservative; therefore, using the glycoprotein gene as the target gene, use the Primer Expre...

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Abstract

The invention relates to the technical field of biology, disclosing a method for fast detecting the reverse transcription-polymerase chain reaction of paralichthys rhabdovirus. The method comprises the following steps of: designing a special primer; extracting RNA of a sample to be detected; having the reverse transcription-polymerase chain reaction; and judging a result by means of agarose gel electrophoresis. The method has the advantages of high speed, good accuracy and specificity and high sensitivity, is suitable for fast testing and quarantining the paralichthys rhabdovirus, and is applicable to aquafarms and disease inspection bureaus, thereby having a wide application prospect.

Description

Technical field: [0001] The invention belongs to the detection and diagnosis technology of aquatic animal pathogens, in particular to the detection method of flounder rhabdovirus. Background technique: [0002] The flounder rhabdovirus (Hirame rhabdovirus, referred to as HRV) virus particle is bullet-shaped, with a size of 80nm×160-180nm, which was first reported in Japan in 1986. This disease has been found in flounder bred in the Yellow Sea, Bohai Sea and other places in my country. The onset season is winter and early spring. When the fish body is onset, the water temperature is generally below 15°C. The disease mainly harms flounder, and artificial infection is highly pathogenic to red sea bream and black sea bream. The virus has also been isolated from sweetfish and unprepared flat carp, and it is also pathogenic to rainbow trout. Fish in the juvenile and larval stages are susceptible to this disease. Diseased fish infected with flounder rhabdovirus have hyperemia or...

Claims

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Application Information

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IPC IPC(8): C12Q1/70C12Q1/68C12R1/93
Inventor 孙颖杰岳志芹刘荭吴斌李叶
Owner 孙颖杰
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