African swine fever polymerase chain reaction (PCR) detection method and oligonucleotide primer pair

An African swine fever, oligonucleotide technology, applied in microorganism-based methods, biochemical equipment and methods, microorganisms, etc., to achieve the effects of suitable length, low cost, high specificity and sensitivity

Active Publication Date: 2014-12-24
QINGDAO AGRI UNIV
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  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

At present, African swine fever has not been introduced into my country, and there is no effective vaccine in the world to prevent and control it. In order to prevent the disease from being introduced into my country and strengthen inspection and quarantine, it is necessary to establish a fast and accurate PCR detection method

Method used

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  • African swine fever polymerase chain reaction (PCR) detection method and oligonucleotide primer pair
  • African swine fever polymerase chain reaction (PCR) detection method and oligonucleotide primer pair
  • African swine fever polymerase chain reaction (PCR) detection method and oligonucleotide primer pair

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Experimental program
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Embodiment

[0016] 1 material

[0017] 1.1 ASFV P72 plasmid

[0018] Using conventional PCR technology, according to the nucleotide sequence of the African swine fever virus P72 gene published in GeneBank, the full-length ASFV P72 gene was amplified and inserted into the pcDNA3.1 vector.

[0019] 1.2 The source and treatment of tissue disease materials

[0020] Swine fever virus (CSFV), porcine reproductive and respiratory syndrome virus (PRRSV), and porcine circovirus type 2 (PCV2) were all purchased by Harbin Veken Biotechnology Development Company.

[0021] The clinical samples were collected from sows with reproductive disorders or piglets with respiratory disorders and aborted fetuses in 14 farms.

[0022] Aseptic treatment of clinical tissue disease material: cut out tissue disease material 2.0g~4.0g, add a small amount of sterilized PBS (containing penicillin and streptomycin), cut into pieces, grind it with a sterile grinder to a paste, then press The volume ratio of 1:4 is diluted with PBS...

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Abstract

The invention belongs to the technical field of biological detection and relates to an African swine fever polymerase chain reaction (PCR) detection method and an oligonucleotide primer pair. The method comprises the following steps: designing a pair of specific primers by referring to a complete genome sequence of 23 ASFVP72 protein strains recorded in GenBank, exploring the optimal reaction system and reaction conditions according to the designed primers, performing sensitivity test, specificity test and reproducibility test on the established PCR reaction method, and performing PCR method detection on clinical samples of sows suffered from reproductive disturbance or piglets with respiratory disturbance symptoms and aborted fetuses from 14 pig farms. The test results prove that the established PCR method can sensitively and rapidly detect the African swine fever virus. The PCR detection method established in the research has high applicability and can be used for detecting the African swine fever disease.

Description

Technical field [0001] The invention relates to a PCR method and an oligonucleotide primer pair for detecting African swine fever, belonging to the technical field of biological detection. Background technique [0002] African swine fever (ASF) is an acute, febrile, highly contagious disease of pigs caused by the African swine fever virus. It has great harm to the pig industry and is an infectious disease closely guarded by various countries. my country classifies it as a type of animal disease. Clinical symptoms and pathological changes are manifested as high fever, skin hemorrhage, respiratory disorders, neurological symptoms, miscarriage, edema and organ hemorrhage, which are similar to acute swine fever. It is easy to be misdiagnosed at the time of diagnosis, and the course of the disease is short and the fatality rate is high, up to 100%. At present, the African swine fever disease has not been introduced into my country, and there is no effective vaccine to prevent and co...

Claims

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Application Information

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Patent Type & AuthorityPatents(China)
IPC IPC(8): C12Q1/70C12Q1/68C12N15/11C12R1/93
Inventor单虎张传美秦晓冰王述柏黄娟杨瑞梅陈琳琳
OwnerQINGDAO AGRI UNIV