General-purpose capripoxvirus TaqMan-MGB probe real-time PCR (polymerase chain reaction) detection method

A technology of sheep pox virus and real-time fluorescence, applied in the field of inspection and quarantine, can solve the problems of low specificity and sensitivity, laborious, time-consuming sheep pox, etc., and achieve the effects of shortening the detection time, simplifying the diagnosis method, and enhancing the specificity

Inactive Publication Date: 2012-07-11
ANIMAL & PLANT & FOOD INSPECTION CENT OF TIANJIN ENTRY EXIT INSPECTION & QUARANTINE BUREAU
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Problems solved by technology

There are many methods for detecting sheeppox virus that have been established, such as virus culture, electron microscope observation, agar diffusion, indirect fluorescent antibody staining, serum neutralization test, enzyme-linked immunosorbent assay technology, etc., but conventional methods are used to detect the presence of sheeppox virus. Disadvantages such as time-consuming, laborious, low specificity and sensitivity

Method used

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  • General-purpose capripoxvirus TaqMan-MGB probe real-time PCR (polymerase chain reaction) detection method
  • General-purpose capripoxvirus TaqMan-MGB probe real-time PCR (polymerase chain reaction) detection method
  • General-purpose capripoxvirus TaqMan-MGB probe real-time PCR (polymerase chain reaction) detection method

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Embodiment

[0046] 1. Extraction of sample genomic DNA

[0047] 1) Treatment Immerse the diseased sheep skin tissue with PBS solution, grind it with a grinder to make a tissue suspension of 1:10, freeze and thaw repeatedly three times, then centrifuge at 600g for 10 min, and take the supernatant for later use.

[0048] 2) Extraction of genomic DNA

[0049] Using TIANGEN company's blood / cell / tissue genomic DNA extraction kit, operate as follows:

[0050] (1) Take 200 μL of the supernatant obtained in step 1), add proteinase K solution, and mix well.

[0051] (2) Add 200 μL buffer GB, mix thoroughly by inversion, and place at 56°C for 10 minutes, during which time, invert and mix several times until the solution becomes clear.

[0052] (3) Add 200 μL of absolute ethanol and mix thoroughly by inversion.

[0053] (4) Add the solution obtained in the previous step and possible flocculent precipitates into an adsorption column CB3 (the adsorption column CB3 is placed in a collection tube), c...

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Abstract

The invention discloses a general-purpose capripoxvirus TaqMan-MGB probe real-time PCR (polymerase chain reaction) detection method and a kit. Through the adoption of the method, the defects in the existing method can be overcome, the capripoxvirus can be diagnosed quickly, accurately and conveniently; emergency control and prevention of goatpox can be facilitated; the economic loss can be reduced; and the method has a very high practical value.

Description

technical field [0001] The invention relates to a PCR detection method for sheep pox virus, in particular to a fluorescent PCR detection method for general-purpose TaqMan-MGB probe of sheep pox virus, belonging to the technical field of inspection and quarantine. Background technique [0002] Goat pox is the collective designation of sheep pox and goat pox, which is an acute, febrile disease of sheep caused by sheep pox virus (SPPV) or goat pox virus (GTPV) of the genus Capripoxvirus. 1. Contact infectious diseases, mainly manifested as fever, papules and herpes on the skin and mucous membranes of hairless or less hairy parts, is the most serious of all animal pox diseases, has a high mortality rate, and can cause huge economic losses. Goatpox is endemic in north-central Africa, central and southwestern Asia, and most of India. The disease occurs in my country's Guizhou, Guangxi, Gansu, Heilongjiang and other places. The World Organization for Animal Health (OIE) lists the...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/70C12Q1/68C12R1/93
Inventor 赵祥平王建华李宁贺艳郑文杰刘伟
Owner ANIMAL & PLANT & FOOD INSPECTION CENT OF TIANJIN ENTRY EXIT INSPECTION & QUARANTINE BUREAU
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