Infectious cDNA clone of canine distemper virus cdv-3 strain and its construction method and application

A technology of CDV-3 and canine distemper virus, which is applied in the direction of recombinant DNA technology, viruses, applications, etc., can solve problems such as threats to human public health safety and economic losses of fur animals

Active Publication Date: 2021-08-03
INST OF SPECIAL ANIMAL & PLANT SCI OF CAAS
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

This not only caused huge economic losses to pet dog and cat breeding and fur animal breeding, but also seriously threatened the safety of human public health

Method used

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  • Infectious cDNA clone of canine distemper virus cdv-3 strain and its construction method and application
  • Infectious cDNA clone of canine distemper virus cdv-3 strain and its construction method and application
  • Infectious cDNA clone of canine distemper virus cdv-3 strain and its construction method and application

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0032] Example 1 Construction of canine distemper virus CDV-3 strain infectious cDNA clone and virus rescue

[0033] 1 Materials and methods

[0034] 1.1 Materials

[0035] 1.1.1 Virus strains and cells

[0036] The CDV-3 strain of canine distemper virus was preserved by the Animal Disease Research Office of the Institute of Special Products, Chinese Academy of Agricultural Sciences; the 293T and Vero cells were preserved by the disease prevention and control team of the Institute of Special Products, Chinese Academy of Agricultural Sciences;

[0037] 1.1.2 Main reagents

[0038] Culture medium DMEM, penicillin streptomycin and calf serum FBS were all purchased from Gibco; eukaryotic expression vector pcDNA3.1 was purchased from Invitrogen; cloning vector pEASY-Blunt or pEASY-Simple-Blunt, high-fidelity enzymes Trans StartFastPfu DNA Polymerase and FITC-labeled goat anti-mouse secondary antibody was purchased from Beijing Quanshijin; various restriction endonucleases and T4...

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Abstract

The invention discloses an infectious cDNA clone of canine distemper virus CDV‑3 strain, a construction method and application thereof. In the present invention, the canine distemper virus CDV-3 strain is used as a template, and the full length of CDV-3 is divided into five fragments for RT-PCR amplification. After enzyme digestion and splicing, the 5 fragments were sequentially inserted into the eukaryotic vector, and the sequences of hammerhead ribozyme and hepatitis D ribozyme were added to the head of F1 and the end of F5, respectively, to obtain the infection of canine distemper virus CDV‑3 strain sex cDNA clones. Then, construct three helper plasmids expressing CDV-3N, P, L protein. The infectious cDNA clone of canine distemper virus CDV‑3 strain and three helper plasmids were co-transfected into 293T cells to obtain rescued recombinant CDV‑3 virus (rCDV‑3). The study found that rCDV‑3 viral titers were obtained up to 10 7.667 TCID 50 / mL, 10 times higher than wtCDV‑3. After infection of Vero cells with rCDV‑3, the highest virus titer was rapidly reached at 36 hours after infection, while the virus content of wtCDV‑3 reached the highest at 72 hours after infection. The proposal of the invention lays a foundation for the development of a novel vaccine against canine distemper virus and the study of its pathogenic mechanism.

Description

technical field [0001] The invention relates to an infectious cDNA clone and its construction method and application, in particular to an infectious cDNA clone of canine distemper virus CDV-3 strain and its construction method and application. The invention belongs to the field of biotechnology. Background technique [0002] Canine distemper (CD) is a highly contagious disease caused by canine distemper virus (CDV) infecting dogs, minks and other carnivorous animals. The disease is highly contagious, has a high incidence rate, and has various clinical symptoms. It is easy to form secondary and mixed infections in the late stage of infection. In recent years, the host of natural infection of canine distemper virus has been expanding continuously. Outbreaks of canine distemper have been reported in wild animals such as tigers, lions, leopards and seals, and even giant panda populations. In January 2015, five giant pandas raised in captivity at the Rare Wildlife Rescue and Br...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N15/85C12N15/45C12N7/01
CPCC07K14/005C12N7/00C12N15/85C12N2760/18421C12N2760/18422
Inventor 薛向红闫喜军卜研陈杰赵建军赵传芳胡博
Owner INST OF SPECIAL ANIMAL & PLANT SCI OF CAAS
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