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Detection reagent and kit for African swine fever virus and application of detection reagent

A technology of African swine fever virus and detection reagents, applied in the field of molecular biology and immunology, can solve the problems of large error in detection results, complicated operation, susceptibility to infection, etc., and achieve the effect of improving accuracy

Pending Publication Date: 2019-01-04
NANJING AGRICULTURAL UNIVERSITY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Existing detection methods have disadvantages such as complex operation, easy to be affected by the environment, easy to infect, unsuitable for early diagnosis, and large error in detection results.

Method used

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  • Detection reagent and kit for African swine fever virus and application of detection reagent
  • Detection reagent and kit for African swine fever virus and application of detection reagent
  • Detection reagent and kit for African swine fever virus and application of detection reagent

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0084] 1 Materials and methods

[0085] 1.1 Materials

[0086] Plasmid DNA of classical swine fever virus and other viruses.

[0087] 1.2 Primer design

[0088] 1.3PCR amplification system is shown in Table 1-1:

[0089] Table 1-1 is the PCR amplification system in embodiment 1

[0090]

[0091]

[0092]The reaction conditions are shown in Table 1-2:

[0093] Table 1-2 is the reaction condition of PCR amplification in embodiment 1

[0094]

[0095] 1.4 Specificity Verification

[0096] Utilize above-mentioned (1.3) PCR reaction system to 1. avian influenza virus; 2. foot-and-mouth disease virus; 3. blue ear virus; 4. porcine chain virus; 5. pseudorabies virus; 6. staphylococcus aureus; 7. salmonella; 8. Escherichia coli; 9. Listeria monocytogenes; 10. Shigella; 11. Vibrio parahaemolyticus; 12. Campylobacter; 13. Hemolytic streptococcus; 14. Streptococcus pneumoniae; 16. Classical swine fever virus; 17. The specificity of negative control (water) was verified by...

Embodiment 2

[0101] To test the sensitivity of the nucleic acid test strip detection method, dilute the PCR template to 1pg, 0.1pg, 0.01pg, 1fg, 0.1fg, 0.01fg, 0.001fg equivalents, perform amplification according to the above (1.3) PCR system, and make 2 parallel.

[0102] 1 Materials and methods

[0103] 1.1 Materials Hog fever virus plasmid DNA

[0104] 1.2 Primer design

[0105] 1.3PCR amplification system is shown in Table 2-1:

[0106] Table 2-1 is the PCR amplification system in embodiment 2

[0107]

[0108] The reaction conditions are shown in Table 2-2:

[0109] Table 2-2 is the reaction condition of PCR amplification in embodiment 1

[0110]

[0111] Take 6 μL of PCR product, add 54 μL of diluent, mix well, add vertically and slowly into the sample hole of the test strip, leave it at room temperature for 3-5 minutes, and read the result with naked eyes; take another 5 μL of PCR product in 1.5% ethidium bromide Electrophoresis was performed on agarose gel electrophores...

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Abstract

The invention discloses a detection reagent and kit for an African swine fever virus and application of the detection reagent. The detection reagent for the African swine fever virus comprises a primer pair, comprising upstream primer and a downstream primer which have sequences as shown by SEQ ID NO: 1 and SEQ ID NO: 2; the end 5' of the upstream primer is labeled with a first nucleic acid marker, and the end 5' of the downstream primer is labeled with a second nucleic acid marker. The method for detecting the African swine fever virus, provided by the invention, is to amplify a gene segmentin a proper length by a labeled specific primer to guarantee high specificity of detection, and the accuracy of a detection result is improved; furthermore, the detection method provided by the invention avoids processes of diluting an amplification product, hybridizing probes and the like and maintains the advantages of intuitiveness, quickness, convenience, maturation, low cost and the like of an immune colloidal gold test strip.

Description

technical field [0001] The invention particularly relates to a detection reagent, kit and application of African swine fever virus, belonging to the technical fields of molecular biology and immunology. Background technique [0002] African swine fever (ASF) is a highly contagious disease of pigs, with a fatality rate of up to 100%. It is caused and spread by African swine fever virus (ASFV). The onset of the disease was very rapid, and it swept from Africa to almost the whole world in less than a century. In view of the serious harm of the disease, the World Health Organization (OIE) listed it as a legally notifiable animal disease, and my country listed it as a first-class animal disease. At present, there is no effective commercial vaccine for the prevention and control of ASFV. On August 1, 2018, the National Center for Alien Animal Disease Research confirmed the first case of ASF in my country in Shenbei District, Shenyang City, and then occurred in Zhengzhou, Henan, ...

Claims

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Application Information

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IPC IPC(8): C12Q1/70C12Q1/6804G01N33/569C12R1/93
CPCC12Q1/6804C12Q1/70G01N33/56983C12Q2531/113
Inventor 薛峰吴晓东戴建君曾德新王志亮苏静任建鸾陈伟
Owner NANJING AGRICULTURAL UNIVERSITY
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