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Separated pathogenicYCCV (yellow catfish calicivirus) as well as specific sequence and application thereof

A technology of calicivirus and yellow catfish, applied to viruses, antiviral agents, virus antigen components, etc., can solve the problems of no homology, etc., and achieve the effects of easy analysis and judgment, improved efficiency, high efficiency and accurate diagnosis

Active Publication Date: 2021-08-06
YANGTZE RIVER FISHERIES RES INST CHINESE ACAD OF FISHERY SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0005] As of the filing date, the pathogen of this virus has been discovered in fish for the first time, and it has no homology with other members of the family Caliciviridae at the nucleotide level, and only about 30% homology at the amino acid level. The study of viral pathogens is still blank

Method used

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  • Separated pathogenicYCCV (yellow catfish calicivirus) as well as specific sequence and application thereof
  • Separated pathogenicYCCV (yellow catfish calicivirus) as well as specific sequence and application thereof
  • Separated pathogenicYCCV (yellow catfish calicivirus) as well as specific sequence and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0035] Discovery of yellow catfish calicivirus and acquisition of specific sequences:

[0036] The applicant collected dying yellow catfish suffering from hemorrhagic disease from different yellow catfish farms for identification of the pathogen. The results showed necrosis and hemorrhage of the spleen and kidney cells of the sick yellow catfish. Ultramicrosection observation results show that there are a large number of spherical non-enveloped virus-like particles in the cells of the visceral tissue of the diseased yellow catfish, with a diameter of about 30-40nm, in the liver, spleen, kidney, heart, gills and other tissues of the yellow catfish. distribution, especially in the spleen and kidney tissues, a large number of viruses multiply. After high-throughput sequencing, it is found that the virus pathogen is a new virus pathogen. The genome sequence is quite different from other members of the Caliciviridae, but the evolutionary relationship It is similar, so it is named ...

Embodiment 2

[0041] Application of the primers designed for the specific sequence of YCCV-NS in the preparation of yellow catfish calicivirus (Yellow catfishcalicivrus, YCCV) detection kit:

[0042] The primers designed for the sequence shown in SEQ ID NO.1 are: YCCV-NS-F: 5'-CGCCTAAAGTTTCCTCCTTGTTGG-3' and YCCV-NS-R: 5'-AAAGTTGGGTGGATTGGTTTGTCAT-3'.

[0043] The amplification system is:

[0044] 1 μl (0.1 μg) of cDNA of the sample to be tested, 1 μl of 10 μM each primer YCCV-NS-F / YCCV-NS-R; 2 μl of 10 mM dNTP Mix; 5 μl of 10×TranStart Taq Buffer (Mg 2+ ) and 1 μl TranStart Taq DNA Polymerase (product of Beijing Quanshijin Biotechnology Co., Ltd.), add ddH2O to make the final volume 50 μl, after mixing, centrifuge for 15s to concentrate the reaction components at the bottom of the tube.

[0045] PCR amplification was performed, and the cycle program was: pre-denaturation at 94°C for 5 min, denaturation at 94°C for 30 s, annealing at 55°C for 30 s, extension at 72°C for 1.5 min, 32 cycles,...

Embodiment 3

[0050] Sensitivity of primers designed for the specific sequence of YCCV-NS:

[0051] The yellow catfish calicivirus YCCV (Yellow catfish calicivrus, YCCV) genome cDNA was serially diluted 10 times: stock solution 0.1ng / μL, 10- 1 ...to 10- 4 , as template cDNA.

[0052] 2) Carry out RT-PCR amplification according to the method in embodiment 2;

[0053] 4) The detection limit of this primer to YCCV can reach 0.1ng / μL ( Figure 4 ).

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Abstract

The invention belongs to the field of virology, fishery biotechnology and aquatic animal medicine, and particularly relates to separated pathogenic YCCV (yellow catfish calicivirus) as well as a specific sequence and application of the separated pathogenic YCCV. The whole genome of the YCCV is shown as SEQ ID NO.2, the specific sequence of the YCCV is shown as SEQ ID NO.1, aiming at the sequence, the applicant designs detection primers YCCC-NS-F and YCCC-NS-R, and when the primers are amplified to a nucleic acid fragment with the size of 523 bp, a patient can be diagnosed as being infected by the YCCV. Gene sequences and primers are particularly suitable for early diagnosis and molecular epidemiological investigation of cultured pelteobagrus fulvidraco with hemorrhagic disease, and are also suitable for monitoring and early warning of caliciviruses of different aquatic animals.

Description

technical field [0001] The invention belongs to the fields of virology, fishery biotechnology and aquatic animal medicine, and more specifically relates to isolated pathogenic yellow catfish calicivirus and its specific sequence and application. Background technique [0002] The yellow catfish Pelteobagrus fulvidraco has delicate meat, rich nutrition, and various breeding methods (monoculture, polyculture, and cage culture). It is a carnivorous, carnivorous, omnivorous, small, famous, and high-quality economically farmed fish widely cultivated in my country [Li Mingde. Beijing: Ocean Press, 2011, 116-122]. The aquaculture industry of yellow catfish in my country has developed rapidly. In recent years, most researches on diseases of yellow catfish have focused on pathogens such as bacteria, fungi and parasites [Zhou Yong et al. China Fishery Quality and Standards, 2019, 9(1): 18- 26; Ye SG, et al. Aquaculture, 2009, 292:6-10]. From March to April 2020, outbreaks of diseases o...

Claims

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Application Information

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IPC IPC(8): C12N7/00C12N15/11A61K39/12A61P31/14C12Q1/70C12Q1/686C12R1/93
CPCA61K39/12A61K2039/552A61P31/14C12N7/00C12N2770/16021C12N2770/16034C12Q1/686C12Q1/701C12Q2565/125
Inventor 刘文枝范玉顶周勇薛明洋李逸群江南孟彦曾令兵
Owner YANGTZE RIVER FISHERIES RES INST CHINESE ACAD OF FISHERY SCI
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