A multiplex PCR kit for the simultaneous detection of six viruses in sheep and goats

A kit and multiple technologies, applied in the direction of biochemical equipment and methods, microbiological determination/inspection, etc., can solve the problems of clinical diagnosis that is not easy to distinguish, lamb growth retardation, economic loss of sheep industry, etc., and achieve short detection time , The detection process is simple and the effect of high sensitivity

Inactive Publication Date: 2019-04-05
NORTHWEST A & F UNIV
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  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

The disease is a common disease, which can cause growth retardation and weight loss of lambs, and cause large economic losses to the sheep industry.
[0009] Bluetongue disease virus, foot-and-mouth disease virus, Peste des petits ruminants virus, sheep pox virus, goat pox virus and sheep mouth disease virus can infect sheep and goats. It manifests as mental depression, elevated body temperature, vesicles and ulcers, mucosal edema and erosion in the oral mucosa and nipples, etc., and the clinical symptoms are very similar, and it is not easy to distinguish them in clinical diagnosis.
At present, the diagnosis of these six epidemic diseases mostly adopts pathogen isolation and identification and conventional serological methods, which takes a long time, and ordinary PCR detection requires different reaction systems, which is time-consuming and laborious

Method used

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  • A multiplex PCR kit for the simultaneous detection of six viruses in sheep and goats
  • A multiplex PCR kit for the simultaneous detection of six viruses in sheep and goats
  • A multiplex PCR kit for the simultaneous detection of six viruses in sheep and goats

Examples

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Effect test

Embodiment 1

[0027] Example 1: Multiplex PCR kit for simultaneous detection of six viruses carried by sheep and goats

[0028] The kit includes six pairs of specific primers, positive control, negative control, reverse transcription solution, PCR mixture and deionized water. Among them, the primers used for the detection of bluetongue virus were BTV-F and BTV-R; the primers used for the detection of foot-and-mouth disease virus were FMDV-F and FMDV-R; the primers used for the detection of Peste des petits ruminants virus were PPRV -F and PPRV-R; the primers used for sheep pox virus detection are SPPV-F and SPPV-R respectively; the primers used for goat pox virus detection are GTPV-F and GTPV-R respectively; the primers used for sheep pox virus detection The primers are ORFV-F and ORFV-R respectively;

[0029] The primers used for bluetongue virus detection are designed for the NS3 gene of bluetongue virus, and the primer sequences are BTV-F and BTV-R, respectively.

[0030] BTV-F: 5'-AGT...

Embodiment 2

[0048] Example 2: Rapid detection of bluetongue virus, foot-and-mouth disease virus, Peste des Petits Ruminants virus, sheep pox virus, goat pox virus and sheep aphthous disease virus

[0049] 1. Materials

[0050] (1) Positive\negative control

[0051] Positive controls: plasmids carrying the NS3 gene of bluetongue virus, plasmids carrying the 3D gene of foot-and-mouth disease virus, plasmids carrying the N gene of the Peste des petits ruminants vaccine strain, plasmids carrying the 119555-119711 segment of the whole gene of sheep pox virus, and plasmids carrying the whole gene of goat pox virus The plasmids of the 110977-111368 segment of the gene and the plasmid mixture carrying the 011 gene of the oropharynx virus were constructed and preserved by the Veterinary Microbiology Laboratory of the School of Veterinary Medicine, Northwest Agriculture and Forestry University. Ltd Synthetics.

[0052] Negative control: deionized water without PPRV, BTV, FMDV, SPPV, GTPV, and ORF...

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Abstract

A multiplex-PCR (polymerase chain reaction) kit for detecting six viruses of sheep and goats simultaneously comprises six pairs of specific primers, wherein the primers used for detection of the bluetongue virus are BTV-F and BTV-R respectively; the primers for detection of the foot and mouth disease virus are FMDV-F and FMDV-R respectively; the primers for detection of the peste des petits ruminants virus are PPRV-F and PPRV-R respectively; the primers for detection of the sheep pox virus are SPPV-F and SPPV-R respectively; the primers for detection of the goat pox virus are GTPV-F and GTPV-R respectively; the primers for detection of the sore mouth disease virus are ORFV-F and ORFV-R respectively. The sequences of the primers are nucleotide sequences shown in SEQ ID NO: 1-12. The multiplex-PCR kit can be used for detecting nucleic acid of the six viruses including the bluetongue virus, the foot and mouth disease virus, the peste des petits ruminants virus, the sheep pox virus, the goat pox virus and the sore mouth disease virus in sheep and goat clinical samples and has the characteristics of high sensitivity, high specificity, simple operation and reduction of detection time.

Description

technical field [0001] The invention relates to a multiplex PCR kit for simultaneously detecting six viruses carried by sheep and goats, in particular to a simultaneous detection of sheep and goat bluetongue virus, foot-and-mouth disease virus, Peste des Petits Ruminants virus, sheep pox virus, goat Multiplex PCR detection kit for poxvirus and aphthus virus. Background technique [0002] Multiplex PCR (multiplex PCR) is an improvement on the basis of ordinary PCR. Multiple pairs of specific primers are added to a PCR reaction system to amplify multiple target fragments for multiple DNA templates or different regions of the same template. Multiplex PCR is mainly used for the simultaneous detection or identification of multiple pathogenic microorganisms and the typing and identification of pathogenic microorganisms, some genetic diseases and cancer genes. Its characteristics are: high efficiency, multiple pathogenic microorganisms can be detected simultaneously in the same PCR...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12Q1/70C12Q1/686
CPCC12Q1/686C12Q1/701C12Q2600/16C12Q2537/143
Inventor 付明哲许信刚何亚鹏张琪鲍长磊
Owner NORTHWEST A & F UNIV
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