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

Method for detecting pseudorabies virus

A technique for pseudorabies virus and detection method, which is applied in the field of molecular biology and can solve the problems of strict annealing temperature, inability to identify PRV, and long time consumption.

Pending Publication Date: 2021-06-04
GUANGXI VETERINARY RES INST
View PDF2 Cites 0 Cited by
  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

For example, Zhang Yueyong et al. (Establishment of a nano-PCR detection method for porcine pseudorabies virus [J]. China Animal Quarantine, 2017, 034(003): 102-105) designed a pair of primers for the PRV gB gene and established a nano-PCR for the detection of PRV method, but since this method can detect both wild strains of PRV and vaccine strains, it is impossible to identify whether the infection of PRV comes from wild strains when used clinically
Ma Xingjie (Establishment and Evaluation of Nano-PCR and Fluorescent Quantitative PCR Methods for Discriminating Porcine Pseudorabies Virus gE / gB / gG [D]. Chinese Academy of Agricultural Sciences, 2014) established triple nano-PCR for the gE / gB / gG gene of PRV, However, this method takes 90 minutes to complete the PCR reaction, which has the disadvantages of long time consumption and strict annealing temperature.

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
  • Method for detecting pseudorabies virus
  • Method for detecting pseudorabies virus
  • Method for detecting pseudorabies virus

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0021] The materials and reagents used in the following examples can be obtained from commercial sources unless otherwise specified. Embodiment 1, screening and optimization of PRV detection method

[0022] 1 Materials and methods

[0023] 1.1 Virus strains and sources of positive cDNA

[0024] Chicken infectious anemia virus GXC060821 strain, porcine pseudorabies virus PRV and chicken astrovirus type 1 positive cDNA (derived from clinical samples) were preserved by Guangxi Veterinary Research Institute; CIAV and PRV were extracted according to the instructions in EasyPure Viral DNA / RNA Kit Viral DNA; DNA and Chicken Astrovirus Type 1 positive cDNA template were stored at -20°C for future use.

[0025] 1.2 Main reagents and instruments

[0026] The general amplification nano-PCR kit was purchased from Shanghai Huzheng Biotechnology Co., Ltd., the PCR instrument was purchased from Bio-rad, USA; the DNA / RNA co-extraction kit EasyPure Viral DNA / RNA Kit was purchased from Beiji...

Embodiment 2

[0061] Embodiment 2, a kind of detection method of PRV

[0062] The method comprises the steps of:

[0063](1) Preparation of template DNA

[0064] The DNA of the PRV virus can be extracted according to the instructions in the EasyPure Viral DNA / RNA Kit.

[0065] (2) Nano-PCR reaction

[0066] Reaction system (20 μL): 1 μL cDNA / DNA template, 10 μL 2×Nano-QPCR buffer, 0.1 μL upstream and downstream primer mixture with a working concentration of 20 μmol / L (the final concentration of primers is 0.1 μmol / L), taq enzyme Mix ( 5U / uL) 0.32μL, add ddH 2 O to make up 20 μL.

[0067] The primer combination is PRV-1F / PRV-1R.

[0068] PCR reaction program: 95°C for 3min; 94°C for 5s, annealing temperature at 50°C for 5s, 72°C for 30s, 20 cycles; 72°C for 5min, 12°C to end the reaction.

[0069] (3) Detection of PCR products

[0070] The amplification results can be verified by 1.2% agarose gel electrophoresis.

Embodiment 3

[0071] Embodiment 3, a kind of detection method of PRV

[0072] The method comprises the steps of:

[0073] (1) Preparation of template DNA

[0074] The DNA of the PRV virus can be extracted according to the instructions in the EasyPure Viral DNA / RNA Kit.

[0075] (2) Nano-PCR reaction

[0076] Reaction system (20 μL): 1 μL cDNA / DNA template, 10 μL 2×Nano-QPCR buffer, 1 μL upstream and downstream primer mixture with a working concentration of 20 μmol / L (the final concentration of primers is 1 μmol / L), taq enzyme Mix (5 U / L uL)0.32μL, add ddH 2 O to make up 20 μL.

[0077] The primer combination is PRV-2F / PRV-2R.

[0078] PCR reaction program: 95°C for 3min; 94°C for 5s, annealing temperature at 50°C for 5s, 72°C for 30s, 20 cycles; 72°C for 5min, 12°C to end the reaction.

[0079] (3) Detection of PCR products

[0080] The amplification results can be verified by 1.2% agarose gel electrophoresis.

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 method for detecting pseudorabies virus. The method is characterized in that PRV detection is carried out by using nano PCR, and the detection is carried out by adopting nucleotide sequences shown in SEQ ID No. 1-2 in a sequence table, the concentration of upstream and downstream primers being 0.1 micromole / L, and the annealing temperature being 50 DEG C, and nucleotide sequences shown in SEQ ID No. 3-4, the concentration of upstream and downstream primers being 1 micromole / L, and the annealing temperature being 50 DEG C. The method has the advantages of high sensitivity, high specificity, loose amplification conditions, specific detection of wild strains and the like.

Description

technical field [0001] The present application relates to the technical field of molecular biology, in particular, to a method for detecting pseudorabies virus. Background technique [0002] Pseudorabies virus (PRV) belongs to the family Herpesviridae (Herpesviridae) α-herpesvirus subfamily (Alphaerpesvirinae) Varicellovirus (Varicellovirus), can infect pigs, sheep, dogs, cattle, cats, rabbits and mice, etc. Animals, which mainly cause symptoms such as fever, itching (except pigs) and encephalomyelitis. [0003] Nano-PCR (nano-PCR) technology is to add nanomaterials (such as gold nanoparticles, silver nanoparticles, carbon nanoparticles and zinc oxide nanoparticles, etc.) To achieve special effects, it provides new ideas and technologies for optimizing conventional PCR. In 2005, Li H et al. (NanoparticlePCR: Nanogold-assisted PCR with enhanced specificity [J] Angewandte chemie-international edition. 2005, 44:2-5) found that gold nanoparticles can improve the activity and a...

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
IPC IPC(8): C12Q1/70C12Q1/686C12R1/93
CPCC12Q1/705C12Q1/686C12Q2565/125C12Q2563/155
Inventor 谢芝勋张民秀张艳芳谢丽基谢志勤李孟邓显文罗思思范晴曾婷婷黄娇玲王盛李丹刘加波
Owner GUANGXI VETERINARY RES INST
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