African swine fever virus attenuated strain lacking MGF360-9L gene and application of African swine fever virus attenuated strain
A technology of African swine fever virus and attenuated strains, applied in the direction of viruses, viral peptides, antiviral agents, etc., can solve the problems of different effects, elevated body temperature, and loss of immunogenicity or protective effect of weak immune side effects strains, etc. Achieving good safety effect
- Summary
- Abstract
- Description
- Claims
- Application Information
AI Technical Summary
Problems solved by technology
Method used
Image
Examples
Example Embodiment
[0038] Example 1 Construction, purification and identification of MGF360-9L gene deletion African swine fever virus
[0039] 1.1 Construction of eGFP screening expression cassette
[0040] The p72 promoter sequence was amplified by PCR, that is, from -97 nt upstream of the p72 gene to the sequence before the ATG start codon; at the same time, using the peGFP-N1 vector as a template, the amplified green fluorescent protein (enhanced green fluorescent protein, eGFP) gene. The two genes were connected by fusion PCR method to obtain the eGFP screening expression cassette gene fragment, named p72-eGFP-SV40 polyA, and the expression cassette sequence contained the SV40 polyA termination sequence.
[0041] 1.2 Construction of homologous recombination transfer vector
[0042] Using pUC57 vector as a backbone vector, a homologous recombination transfer vector for MGF-360-9L gene knockout was constructed. The specific steps are as follows: Design 1.5 kb of the upstream and downstream sequence...
Example Embodiment
[0047] Example 2 Titration of virus titer
[0048] The titration of African swine fever virus adopts 50% haemadsorption (HAD) 50 ) Method to operate. HAD 50 The trial was based on the literature (Borca MV, Ramirez-Medina E, Silva E, Vuono E, Rai A, Pruitt S, Holinka LG, Velazquez-Salinas L, Zhu J, GladueDP. Development of a highly effective African swine fever virus vaccine by deletion of the I177Lgene results in sterile immunity against the current epidemic Eurasiastrain.JVirol. 2020. pii: JVI.02017-19) and make appropriate adjustments: inoculate primary PBMC in a 96-well cell culture plate, and 10 times the sample to be tested Inoculate 0.02ml in each well. Virus infection can be judged according to the rosette formed by red blood cells gathering around the infected cells. Observation for 7 days, according to Reed and Muench method (Reed, L. and H. Muench, A simple method of estimaing fifty percent endpoints. American Journal of Epidemiology 1938.27: p.493-497) calculates the ...
Example Embodiment
[0050] Example 3 Animal challenge experiment
[0051] In order to detect the pathogenicity of the gene knockout ΔMGF-360-9L strain, this example uses 10 HAD 50 The dose was injected intramuscularly into piglets to evaluate its pathogenicity. Specifically, in this example, 10 healthy Landrace piglets negative for the African swine fever antigen antibody were divided into 2 groups, each of five groups was used to attack the parental strain (wild African swine fever virus strain) and the deleted strain respectively. After that, the body temperature changes were measured every day, peripheral blood and saliva were collected, and the blood content of ASFV virus was determined by fluorescent quantitative PCR method. The observation was terminated on 19 days.
[0052] Results: The pathogenicity evaluation showed that the parent strain was injected intramuscularly with 10 HAD 50 Later, the typical symptoms of ASFV appeared: high temperature, and all died in the later period; while the miss...
PUM
Abstract
Description
Claims
Application Information
- R&D Engineer
- R&D Manager
- IP Professional
- Industry Leading Data Capabilities
- Powerful AI technology
- Patent DNA Extraction
Browse by: Latest US Patents, China's latest patents, Technical Efficacy Thesaurus, Application Domain, Technology Topic.
© 2024 PatSnap. All rights reserved.Legal|Privacy policy|Modern Slavery Act Transparency Statement|Sitemap