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Triple fluorescent quantitative PCR primer group for detecting different types of porcine circovirus, kit and application

A porcine circovirus, fluorescence quantitative technology, applied in recombinant DNA technology, microbial determination/inspection, biochemical equipment and methods, etc., can solve the problems of complicated operation, time-consuming and laborious, low sensitivity, etc.

Pending Publication Date: 2021-09-17
浙江省动物疫病预防控制中心
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  • Claims
  • Application Information

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Problems solved by technology

Moreover, conventional pathogenic isolation and serological detection methods have the disadvantages of complex operations, time-consuming and labor-intensive, low sensitivity, and can only detect one type at a time, especially PCV4 is a newly discovered virus. detection of multiple types of porcine circovirus, and no related studies

Method used

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  • Triple fluorescent quantitative PCR primer group for detecting different types of porcine circovirus, kit and application
  • Triple fluorescent quantitative PCR primer group for detecting different types of porcine circovirus, kit and application
  • Triple fluorescent quantitative PCR primer group for detecting different types of porcine circovirus, kit and application

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Experimental program
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Embodiment 1

[0051] 1. Entrust Sangon Biotech (Shanghai) Engineering Co., Ltd. to synthesize and label the primers and probes shown in Table 1.

[0052] 2. Use the viral nucleic acid extraction kit MagNA pure 96 DNA and viral NA Small VoumeKit to extract viral nucleic acid according to the operating instructions, and store the obtained viral nucleic acid in a -80°C refrigerator for later use.

[0053] 3. Preparation of recombinant plasmid standard products of porcine circovirus type 2, porcine circovirus type 3 and porcine circovirus type 4

[0054] According to the target gene fragment detected in the triple fluorescent quantitative detection method of porcine circovirus type 2, porcine circovirus type 3 and porcine circovirus type 4, primers (Table 2) were designed to amplify porcine circovirus type 2 respectively 1. The nucleic acid fragments of porcine circovirus type 3 and porcine circovirus type 4 detection genes, the specific operations are as follows: respectively use the above-men...

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Abstract

The invention provides a triple fluorescent quantitative PCR primer group for detecting different types of porcine circovirus, a kit and application, and relates to the technical field of biological detection. According to the triple fluorescent quantitative PCR primer group for detecting different types of porcine circovirus, primers and probes are synthesized according to conserved sequences of porcine circovirus type 2, porcine circovirus type 3 and porcine circovirus type 4 in Genbank, and meanwhile, the kit is further constructed and used for porcine epidemic disease detection. 83 porcine tissue samples from Zhejiang Province are detected by utilizing an established fluorescent quantitative PCR system, and a result shows that the primer group is good in specificity, repeatability and stability, relatively high in consistency and relatively high in accuracy. According to the triple fluorescent quantitative PCR primer group for detecting different types of porcine circovirus, the lowest detection values of PCV2 and PCV3 are both 10 copies / microliter, the lowest detection value of PCV4 is 1 copy / microliter, and the defects of low sensitivity, prone laboratory pollution and the like of the current triple PCR detection method for the porcine circovirus type 2, the porcine circovirus type 3 and the porcine circovirus type 4 are overcome.

Description

technical field [0001] The invention belongs to the technical field of biological detection, and in particular relates to a triple fluorescence quantitative PCR primer set, a kit and an application for detecting different types of porcine circovirus. Background technique [0002] Porcine circovirus (PCV) is a circular single-stranded DNA virus that can replicate autonomously in mammalian cells. Four types of PCV have been found, namely porcine circovirus type 1 (PCV1), porcine circovirus type 2 (PCV2), porcine circovirus type 3 (PCV3) and porcine circovirus type 4 (PCV4). . PCV1 is a virus found in the process of PK15 cell culture, and studies have shown that it is not pathogenic to pigs; PCV2 is pathogenic, mainly infecting domestic pigs and wild boars, and most of them are infected at the age of 4 to 11 weeks; PCV3 and wild boars were first reported in 2016. Myocarditis is related to glomerulonephritis; the Veterinary Virology Research Group of the School of Biology, Hun...

Claims

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Application Information

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IPC IPC(8): C12Q1/70C12Q1/6851C12N15/11
CPCC12Q1/701C12Q1/6851C12Q2600/16
Inventor 谢荣辉徐辉安慧婷赵灵燕王雅婷黄靖刘霞张红丽孙冰冰董建斐冯肖肖张璐陈伟良薛银柴娟虞一聪
Owner 浙江省动物疫病预防控制中心
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