Specific primer, probe and kit for detecting Chinese soft-shelled turtle flavivirus

A Chinese soft-shelled turtle, flavivirus technology, applied in the direction of viruses, viral peptides, viruses/phages, etc., to achieve the effect of high mortality, long incubation period, and conducive to green sustainable development

Active Publication Date: 2020-12-18
ZHEJIANG INST OF FRESH WATER FISHERIES +1
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

There is no report about the pathogenic c

Method used

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  • Specific primer, probe and kit for detecting Chinese soft-shelled turtle flavivirus
  • Specific primer, probe and kit for detecting Chinese soft-shelled turtle flavivirus
  • Specific primer, probe and kit for detecting Chinese soft-shelled turtle flavivirus

Examples

Experimental program
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Effect test

Embodiment 1

[0032] 1. Sample collection and virus purification

[0033] Take out the liver, spleen, lung, kidney and intestinal tissues of the diseased soft-shelled turtle, and homogenize them on ice. The homogenate was centrifuged at 12,000 g for 20 min, the supernatant was taken, filtered through a 0.22 μm bacterial filter, the filtrate was centrifuged at 40,000 g for 3 h, and the precipitate was resuspended in TNE buffer as the virus-containing crude extract. The above virus liquid was purified by discontinuous sucrose density gradient centrifugation.

[0034] 2. Viral nucleic acid extraction and high-throughput sequencing

[0035] Take 200 μL of the purified virus liquid, use the QIAamp MinElute Virus Spin Kit (QIAGEN Germany) kit to extract viral nucleic acid, and send it to Sangon Bioengineering (Shanghai) Co., Ltd. for RNA and DNA library construction, and then separately High-throughput sequencing, in which high-throughput sequencing of RNA library construction, splicing out par...

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PUM

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Abstract

The invention discloses a specific primer, a probe and a kit for detecting Chinese soft-shelled turtle flavivirus, and belongs to the technical field of biological detection. According to the specificprimer, the probe and the kit, a pathogenic factor causing the disease is proved to be a virus by collecting a diseased turtle tissue sample and performing pathogen analysis. After a partial sequenceof the virus is obtained by adopting an RACE technology, a primer group, the probe and the kit for specifically detecting the virus are designed in a targeted manner, a method for sensitively, specifically and quickly detecting the Chinese soft-shelled turtle flavivirus is provided, the blank that no Chinese soft-shelled turtle flavivirus detection method exists at present is filled up, and the method is suitable for definite diagnosis, screening and prevention of the Chinese soft-shelled turtle flavivirus. The specific primer, the probe and the kit provide technical support for fry quarantine, provide a tool for cutting off virus transmission from the source, and are beneficial to green sustainable development of the Chinese soft-shelled turtle breeding industry.

Description

technical field [0001] The invention relates to the technical field of biological detection, in particular to a specific primer, probe and kit for detecting Chinese soft-shelled turtle yellow virus. Background technique [0002] Chinese soft-shelled soft-shelled turtle (Pelodiscus sinensis), commonly known as soft-shelled turtle, tuna, and bastard, belongs to the order Testudines, and the wild soft-shelled soft-shelled turtle is widely distributed in China. Chinese soft-shelled turtle has high nutritional value and can be used as medicine. It is deeply loved by consumers and has always been an important economic animal for aquaculture in my country. In recent years, there have been explosive death cases in some Chinese soft-shelled turtle farms in Zhejiang Province. The main characteristics of the disease are: First, the food intake has decreased. In the early stage, some sick soft-shelled turtles floated on the water or lay on the shore, showing symptoms of weakness and ano...

Claims

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Application Information

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IPC IPC(8): C12N15/40C12N15/63C12Q1/70C12Q1/6851C12N15/11C12R1/93
CPCC07K14/005C12N15/63C12N2770/24122C12Q1/6851C12Q1/701C12Q2531/113C12Q2563/107C12Q2545/113
Inventor 袁雪梅刘莉吕孙建张海琪
Owner ZHEJIANG INST OF FRESH WATER FISHERIES
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