SARS-Cov gene vaccine based on epi-position and its contruction

A gene vaccine and epitope technology, applied in the fields of bioengineering and immunology, can solve the problems of inefficient expression of short peptides, achieve high specificity, good expected clinical value, and overcome poor expression efficiency

Inactive Publication Date: 2005-08-24
FUDAN UNIV
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  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0005] At present, due to the inefficiency of short peptide expression, all reported SARS-Cov vaccines have not effectively induced the body's immune response against the virus based on epitope-based gene vaccines

Method used

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  • SARS-Cov gene vaccine based on epi-position and its contruction
  • SARS-Cov gene vaccine based on epi-position and its contruction
  • SARS-Cov gene vaccine based on epi-position and its contruction

Examples

Experimental program
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Effect test

Embodiment 1

[0058] Example 1 Construction, Identification and Expression of Epitope-Based SARS-Cov Gene Vaccine

[0059] 1. Construction of epitope-based SARS-Cov gene vaccine

[0060]After software prediction combined with network database prediction, the polypeptide sequence as shown in Table 1 was first determined as the target epitope, according to the mammalian codon reported by Kotsopoulou E. (Journal of Virology; Vol.74, No.10, 4839-4852) After preference optimization, the gene fragments shown in Table 2 were artificially synthesized. After the synthetic fragments were phosphorylated by T4 polynucleotide kinase for 1 hour, they were heated and boiled for 5 minutes, kept warm, cooled and annealed slowly, and then connected in pairs to obtain ab, cd, ef, gh (connected at 16°C for 6 hours). After the cassettes were recovered, abcd and efgh were ligated in pairs again, which were recovered and ligated again, and the target gene fragments with BglII and EcoRI restriction sites at both ...

Embodiment 2

[0068] Example 2 The epitope-based SARS-Cov gene vaccine immunizes mice to induce specific humoral immunity

[0069] Experimental Study of Immune Response

[0070] Experimental animals: 6-8 weeks old female BALB / c(H-2 d ) mice, weighing 16-18 grams, were purchased from Shanghai Experimental Animal Center, Chinese Academy of Sciences.

[0071] Experimental grouping and animal immunization: 12 healthy female BALB / c mice each, divided into 2 groups: (1) immunization group with purified pVAON33 empty plasmid vector as control, 6 mice; (2) Purified pVAON33-epis plasmid Immunization group, 6 rats. Intramuscularly immunize female BALB / c mice with mild anesthesia and inject into the tibialis anterior muscle: intraperitoneally inject mice with 0.75% pentobarbital sodium 100-150ul (according to 50ug / g body weight), expose the tibialis anterior muscle, and use a 1ml syringe Insert the needle vertically from the middle of the muscle, insert the needle tip to a depth of ...

Embodiment 3

[0074] Example 3 Epitope-based SARS-Cov gene vaccine immunizes mice to induce specific cells

[0075] Experimental Study of Immune Response

[0076] Experimental animals: 6-8 weeks old female BALB / c(H-2d) mice, weighing 16-18 grams, purchased from Shanghai Experimental Animal Center, Chinese Academy of Sciences.

[0077] Experimental grouping and animal immunization: 12 healthy female BALB / c mice each, divided into 2 groups: (1) immunization group with purified pVAON33 empty plasmid vector as control, 6 mice; (2) Purified pVAON33-epis plasmid Immunization group, 6 rats. Intramuscularly immunize female BALB / c mice with mild anesthesia and inject into the tibialis anterior muscle: intraperitoneally inject mice with 0.75% pentobarbital sodium 100-150ul (according to 50ug / g body weight), expose the tibialis anterior muscle, and use a 1ml syringe Insert the needle vertically from the middle of the muscle, insert the needle tip to a depth of 2-3mm, slowly insert th...

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Abstract

A SARS-Cov gene vaccine based on epitope is configured from the carrier which is a plasmid able to be used for human body and the target antigen which is several B cell epitopes in the extrinsic B protein antigen of human SARS coronavirus through codon optimizing and genetic engineering. Its preparing process is also disclosed.

Description

technical field [0001] The invention belongs to the fields of bioengineering and immunology, and relates to a genetic vaccine. Further, the invention relates to an epitope-based SARS-Cov genetic vaccine constructed by means of genetic engineering. Background technique [0002] A newly isolated coronavirus (SARS-Cov) is currently recognized as the causative agent of the outbreak of atypical pneumonia. It poses a huge threat to human health and causes huge losses to the economy of our country. It has infected more than 8,000 people on five continents and caused a mortality rate greater than 10%. According to the results of genome sequencing analysis, it is currently known that the main structural proteins of the virus include Spike (S), M and E proteins on the membrane surface, and the N protein connected to the RNA genome, which may become the main target for the development of vaccines. As an infectious disease caused by a virus, vaccines are widely considered to be the mos...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): A61K48/00A61P11/00A61P31/14
Inventor 熊思东汪晓华王缨储以微
Owner FUDAN UNIV
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