Looking for breakthrough ideas for innovation challenges? Try Patsnap Eureka!

Primer for rapidly detecting exogenous virus in porcine pseudorabies live vaccine and application of primer in kit

A technology for detecting primers and porcine circovirus, which is applied in the field of molecular biology testing, can solve the problems of wasting detection time, increasing labor load, and reducing detection efficiency, and achieves easy operation, low requirements for personnel levels and laboratory conditions, and improved The effect of detection efficiency

Inactive Publication Date: 2021-06-01
SHANDONG BINZHOU ANIMAL SCI & VETERINARY MEDICINE ACADEMY
View PDF3 Cites 1 Cited by
  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, at present, there are no multiple fluorescent PCR detection kits for simultaneous detection of ASFV, BVDV, and PCV2 at home and abroad, so that the detection of live vaccines needs to be carried out step by step, which wastes detection time and increases labor and reduces detection efficiency. Therefore, it is of great significance to design a kit for simultaneous detection of ASFV, BVDV, and PCV2 to meet the needs of rapid quality control of actual vaccines.

Method used

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
View more

Image

Smart Image Click on the blue labels to locate them in the text.
Viewing Examples
Smart Image
  • Primer for rapidly detecting exogenous virus in porcine pseudorabies live vaccine and application of primer in kit
  • Primer for rapidly detecting exogenous virus in porcine pseudorabies live vaccine and application of primer in kit
  • Primer for rapidly detecting exogenous virus in porcine pseudorabies live vaccine and application of primer in kit

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0043] Establishment of multiplex fluorescent PCR reaction system for African swine fever virus, bovine viral diarrhea virus and porcine circovirus type 2

[0044] 1. Primer design and preparation

[0045] Find the VP72 gene sequence of different strains of African swine fever virus (ASFV), the 5'-UTR gene sequence of bovine virus diarrhea virus (BVDV) and the ORF1 gene sequence of different strains of porcine circovirus type 2 (PCV2) from Gen Bank , through the comparison analysis, select the conserved regions and design a pair of amplification primers, the sequences are as follows:

[0046] The specific primer I used for the amplification of African swine fever virus is:

[0047] SEQ ID NO. 1: upstream primer ATCCAAACAGCAGGTAAA;

[0048] SEQ ID NO.2: downstream primer CTGAGGGAATAGCAAGGT;

[0049] The specific primer II for bovine viral diarrhea virus amplification is:

[0050] SEQ ID NO.3: upstream primer GATGGCCGAATCCCTGAG;

[0051] SEQ ID NO. 4: downstream primer GTTA...

Embodiment 2

[0076] Sensitivity detection

[0077] The positive control substance in Example 1 was serially diluted 10 times, and the concentration of the standard pMD-VP72 of African swine fever virus plasmid in the mixture was 8.2×10 4 copies / μL~8.2×10 -1 copies / μL, the concentration of standard pMD-5' of bovine viral diarrhea virus plasmid is 5.7×10 4 copies / μL~5.7×10 -1 copies / μL, the concentration of porcine circovirus type 2 plasmid standard pMD-ORF1 is 3.6×10 4 copies / μL~3.6×10 -1 copies / μL, each gradient was repeated 3 times, and the multiplex fluorescence quantitative PCR detection was carried out under the guidance of the primers and TaqMan in Example 1. The detection system and detection conditions were as described in Example 1.

[0078] The test results showed that the minimum detection limit of the multiplex fluorescent PCR method for African swine fever virus was 8.2 copies / μL, the minimum detection limit for bovine viral diarrhea virus was 5.7 copies / μL, and the minimum...

Embodiment 3

[0080] specific detection

[0081] Positive control pMD-VP72 / 5' / ORF1, swine fever virus, porcine reproductive and respiratory syndrome virus, porcine pseudorabies virus, porcine epidemic diarrhea virus, porcine transmissible gastroenteritis virus, porcine rotavirus, porcine The delta coronavirus genome was used as a template to conduct a specific test, and nuclease-free water was used as a negative control; nucleic acid extraction, detection system and detection conditions were as described in Example 1.

[0082] The results showed that the melting curve Tm values ​​of the positive control pMD-VP72 / 5' / ORF1 showed melting peaks at 82±0.5°C, 84±0.5°C, and 86±0.5°C, while the other samples and negative controls did not have the above three specific peaks. It is proved that the kit of the present invention has good specificity and no crossover with other common viruses, and can specifically detect exogenous African swine fever virus, bovine viral diarrhea virus and porcine circovi...

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
Login to View More

PUM

No PUM Login to View More

Abstract

The invention provides a primer for rapidly detecting an exogenous virus in a porcine pseudorabies live vaccine and application of a primer in a kit. The primer comprises a specific primer I aiming at an African swine fever virus, a specific primer II aiming at a bovine viral diarrhea virus and a specific primer III aiming at a porcine circovirus type 2; the primers are applied to the kit, and rapid screening and detection of three pathogens, namely, African swine fever virus (ASFV), bovine viral diarrhea virus (BVDV) and porcine circovirus type 2 (PCV2), are simultaneously completed in a reaction system; by arranging the detection primers, the specificity, the stability and the sensitivity for detecting the ASFV, the BVDV and the PCV2 can be improved; when being applied, the kit has the advantages of simplicity in operation, short detection time and no limitation of the level of detection personnel and the level of a laboratory.

Description

technical field [0001] The invention belongs to the technical field of molecular biology testing, and in particular relates to a primer for rapid detection of exogenous virus in a porcine pseudorabies live vaccine and its application in a kit. Background technique [0002] Porcine pseudorabies is an acute viral infectious disease caused by pseudorabies virus (PRV). Up to 100%, it is one of the common and frequently-occurring important diseases of pig herds, which seriously endangers the development of pig breeding industry. Currently, there is no effective treatment for porcine pseudorabies, which is mainly prevented by live vaccination. [0003] The production of porcine pseudorabies virus live vaccine requires a variety of raw and auxiliary materials such as seed toxin, serum, trypsin and cell matrix. These raw and auxiliary materials are prepared from animal-derived substances, which have the risk of contaminating various viruses. biosafety hazard. [0004] At present,...

Claims

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
Login to View More

Application Information

Patent Timeline
no application Login to View More
Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/70C12Q1/686C12N15/11C12R1/93
CPCC12Q1/701C12Q1/686C12Q2600/16
Inventor 于新友王金良沈志强孟卫芹李天芝魏凤董林
Owner SHANDONG BINZHOU ANIMAL SCI & VETERINARY MEDICINE ACADEMY
Who we serve
  • R&D Engineer
  • R&D Manager
  • IP Professional
Why Patsnap Eureka
  • Industry Leading Data Capabilities
  • Powerful AI technology
  • Patent DNA Extraction
Social media
Patsnap Eureka Blog
Learn More
PatSnap group products

Browse by: Latest US Patents, China's latest patents, Technical Efficacy Thesaurus, Application Domain, Technology Topic, Popular Technical Reports.

© 2024 PatSnap. All rights reserved.Legal|Privacy policy|Modern Slavery Act Transparency Statement|Sitemap|About US| Contact US: help@patsnap.com