Subunit vaccine for novel coronavirus and application of subunit vaccine
A coronavirus, a new type of technology, applied in the direction of viruses, viral peptides, antiviral agents, etc., to achieve the effects of improving stability, increasing immunogenicity, and prolonging plasma half-life
- Summary
- Abstract
- Description
- Claims
- Application Information
AI Technical Summary
Problems solved by technology
Method used
Image
Examples
Embodiment 1
[0020] Example 1: Amplification of SARS2-RBD and SARS2-RBD-Fc fragment genes
[0021] According to the reported clinically isolated SARS-CoV-2 virus (GenBank: QHR63250.2) gene sequence, the forward and reverse primers of the SARS2-RBD gene were designed to amplify the target fragment, and the Fc fragment gene of human IgG1 (GenBank: CAC20454. 1) Gene sequence design primers to amplify the Fc fragment and recombine:
[0022] Forward primer:
[0023]
[0024]
[0025] Note: The horizontal line in SEQ No. 3 is the recombined part on the vector.
[0026] Reverse primer:
[0027]
[0028] Note: The horizontal line in SEQ No. 7 is the recombined part on the vector.
[0029] Use forward primers (1) / (2) and reverse primers (1) to SARS-CoV-2 virus and use forward primers (3) and reverse primers (2) to IgG1 Fc fragment gene bases respectively The sequence was used as a template to perform PCR to obtain SARS2-RBD and Fc gene fragments, and the forward primer (1) and reverse primer (2) were used t...
Embodiment 2
[0032] Example 2: Expression and purification of SARS2-RBD-Fc protein
[0033] Expression using mammalian cell 293F expression system. One day before transfection, 293F cells (control cell density of 5×10 5 A / ml) 40mL was inoculated in a 125mL suspension cell culture flask. Dilute 40μg plasmid (pCAGGS-SARS2-RBD-Fc) in 4mL culture medium and mix gently, then dilute 60μL PEI (polyethylenimine catalog number 24765-1) in the culture medium and mix gently. After incubating at room temperature for 20 minutes, add them dropwise to the cells. Put the cells in a suspension incubator, 250 rpm, 37℃ 8% CO 2 Suspension culture.
[0034] After culturing for 120 hours, the culture supernatant was collected and Goat Anti-human-IgG Fc Antibody was used as an antibody to detect the expression of pCAGGS-SARS2-RBD-Fc by Western Blot ( image 3 ).
[0035] After detecting the expression of pCAGGS-SARS2-RBD-Fc, expand the cell culture and transfection scale to express a large amount of receptor binding...
Embodiment 3
[0037] Example 3: Mouse Immunization of SARS2-RBD-Fc Protein
[0038] The fusion proteins SARS2-RBD-Fc and SARS2-RBD were mixed uniformly with Imject Alum adjuvant (ThermoScientific), and the final protein concentration was 1 mg / ml. Female BABL / c mice aged 4-6 weeks were divided into 4 groups with 4 mice in each group. The first group is to immunize mice with SARS2-RBD+adjuvant 20μl / mouse by nasal drip; the second group is to immunize mice with SARS2-RBD-Fc+adjuvant 20μl / mouse by nasal drip; The group received 20 μl / mouse of SARS2-RBD-Fc+adjuvant by intramuscular injection in the leg; the fourth group was immunized with 20 μl / mouse of PBS+adjuvant by nasal drip. Two weeks later, the same dose was used for the second immunization. Two weeks after the second immunization (28 days in total), a certain amount of blood was taken from the mouse orbital venous plexus, and the specific antibody titer in the serum was measured by ELISA (ELISA coated SARS2-RBD protein). The result is F...
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, Popular Technical Reports.
© 2024 PatSnap. All rights reserved.Legal|Privacy policy|Modern Slavery Act Transparency Statement|Sitemap|About US| Contact US: help@patsnap.com