Gene chips and kits for detecting foot-and-mouth disease virus, swine fever virus and porcine reproductive and respiratory syndrome virus

A technology for respiratory syndrome and foot-and-mouth disease virus, which is applied in the directions of microorganism-based methods, biochemical equipment and methods, and microbial determination/inspection, etc. It can solve the problems of stability and sensitivity test, etc., to achieve the effect of good clinical application prospect, rapid detection and high sensitivity.

Active Publication Date: 2016-08-24
SICHUAN AGRI UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0006] At present, the patent with the notification number CN1274849C discloses a gene chip for simultaneous detection of foot-and-mouth disease virus, swine fever virus, pseudorabies virus and porcine reproductive and respiratory syndrome virus. Specific and sensitive simultaneous detection of foot-and-mouth disease virus, swine fever virus and porcine reproductive and respiratory syndrome virus

Method used

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  • Gene chips and kits for detecting foot-and-mouth disease virus, swine fever virus and porcine reproductive and respiratory syndrome virus
  • Gene chips and kits for detecting foot-and-mouth disease virus, swine fever virus and porcine reproductive and respiratory syndrome virus
  • Gene chips and kits for detecting foot-and-mouth disease virus, swine fever virus and porcine reproductive and respiratory syndrome virus

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0073] Embodiment 1 Detection method of the present invention

[0074] 1. Materials and instruments

[0075] The same experimental materials and instruments as mentioned above.

[0076] 2. Experimental method

[0077] 2.1 Design and synthesis of PCR primers and detection probes

[0078] Log in to GenBank to obtain the gene sequences of representative strains of FMDV, CSFV, and PRRSV main genotypes (serotypes), use the MegAlign module of DNAStar software to perform homology analysis, select conserved genes as detection target regions, and use Oligo7.0 The software designs amplification primers and oligonucleotide probes. When labeling samples, the downstream primers are primers modified by Cy3 at the 5' end; the 5' end of the oligonucleotide probe is added with a Poly T15 linker and aminated. The primer sequences are shown in Table 1, and the oligonucleotide probes are shown in Table 2. After Blast comparison, they were sent to Shanghai Sangong for synthesis.

[0079] Tabl...

Embodiment 2

[0165] Embodiment 2 specificity test

[0166] 1. Test method

[0167] Detection of porcine circovirus (PCV-2), porcine parvovirus (PPV), pseudorabies virus (PRV), porcine Japanese encephalitis virus (JEV) and porcine epidemic diarrhea virus (PEDV) by multiplex asymmetric RT-PCR , porcine transmissible gastroenteritis virus (TGEV), bovine viral diarrhea / mucosal disease virus (BVDV) nucleic acid template amplification markers, and the amplified products were detected with the gene chip prepared in Example 1 respectively to evaluate its specificity.

[0168] The method of multiplex RT-PCR amplification is as follows:

[0169] Extract the nucleic acid of the sample to be tested (use the RNA column extraction kit and DNA column extraction kit of Beijing Century Yuanheng Company to extract the nucleic acid of the corresponding virus), multiplex RT-PCR amplification, and the reaction system is: use One Step PrimeScriptTMRT-PCR Kit Amplification kit, Buffer III (2x) 12.5 μL, Enzyme ...

Embodiment 3

[0176] Embodiment 3 sensitivity test

[0177] 1. Test method

[0178] According to the method of Example 1, the recombinant plasmid PMD3D (8.34 × 10 9 copies / μL), PMD-W (1.06×10 9 copies / μL), PMD-C (9.02×10 9 copies / μL), PMDP (3.76×10 9 copies / μL), PMDHP (3.65×10 9 copies / μL) were diluted 10×, amplified by multiple asymmetric RT-PCR, and the amplified products were detected by gene chip to evaluate the sensitivity of the detection gene chip system.

[0179] The multiplex RT-PCR amplification method and gene chip detection method are the same as in Example 2.

[0180] 2. Results

[0181] Adopt the method of the present invention to detect plasmid PMD3D, the minimum detection concentration can reach 8.34 * 10 2 copies / μL, test results see Figure 20 .

[0182] Adopt the method of the present invention to detect plasmid PMD-W, the minimum detection concentration can reach 1.06 * 10 2 copies / μL, test results see Figure 21 .

[0183] Adopt the method of the present inv...

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Abstract

The invention discloses a gene chip and a kit for detecting a foot and mouth disease virus, a hog cholera virus and a porcine reproductive and respiratory syndrome virus. According to the gene chip, the detection kit and the detection method disclosed by the invention, the foot and mouth disease virus, the hog cholera virus and the porcine reproductive and respiratory syndrome virus can be accurately and effectively detected, and the gene chip and kit are high in specificity, high in sensitivity, low in time consumption, rapid in detection and good in application prospects.

Description

technical field [0001] The invention relates to a gene chip and a detection method for detecting foot-and-mouth disease virus, swine fever virus and porcine reproductive and respiratory syndrome virus. Background technique [0002] Foot-and-mouth disease (FMD) is an acute, febrile, and highly contagious disease of cloven-hoofed animals caused by foot-and-mouth disease virus (FMDV), characterized by blisters on the oral mucosa, hooves, breasts, and skin. , followed by a class of extremely fast-spreading infectious diseases of ulcers. The disease seriously endangers animal husbandry production, affects international trade and national reputation, and has received widespread attention from governments of all countries. Animals susceptible to FMD include more than 70 species of domestic and wild artiodactyls, including cattle, sheep, goats, camels, and pigs. The disease has been widely prevalent in the world, causing huge economic losses to the world's aquaculture industry. I...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12Q1/70C12Q1/68C12N15/11C12R1/93
CPCC12Q1/701
Inventor 文心田李金海李兴玉黄小波曹三杰文翼平伍锐
Owner SICHUAN AGRI UNIV
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