A fusion protein, recombinant vector, recombinant dendritic cell and application thereof for transmembrane expression of novel coronavirus antigen s2
A coronavirus and fusion protein technology, applied in the field of whole-cell vaccines, can solve problems such as virus infection, achieve strong immunogenicity, high degree of conservation, and prevent virus invasion
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Embodiment 1
[0056] Plasmid construction:
[0057] Gene synthesis of the fusion gene CD4 signal peptide-S2 protein-flag tag-CD4 transmembrane domain (entrusted to Jinweizhi Biotechnology Co., Ltd.), the specific sequence is:
[0058] atgaaccggggagtcccttttaggcacttgcttctggtgctgcaactggcgctcctcccagcagccactcagggaagctttatcgaggacctgctgttcaacaaggtgaccctggccgatgccggcttcatcaagcagtatggcgattgcctgggcgacatcgcagccagggacctgatctgcgcccagaagtttaatggcctgaccgtgctgccacccctgctgacagatgagatgatcgcacagtacacaagcgccctgctggccggcaccatcacatccggatggaccttcggcgcaggagccgccctccagatcccctttgccatgcagatggcctataggttcaacggcatcggcgtgacccagaatgtgctgtacgagaaccagaagctgatcgccaatcagtttaactccgccatcggcaagatccaggacagcctgtcctctacagccagcgccctgggcaagctccaggatgtggtgaatcagaacgcccaggccctgaataccctggtgaagcagctgagcagcaacttcggcgccatctctagcgtgctgaatgacatcctgagccggctggacaaggtggaggcagaggtgcagatcgaccggctgatcaccggccggctccagagcctccagacctatgtgacacagcagctgatcagggccgccgagatcagggccagcgccaatctggcagcaaccaagatgtccgagtgcgtgctgggccagtctaagagagtggacttttgtggcaagggctatc...
Embodiment 2
[0090] Material:
[0091] Cells: DC cells
[0092] Medium: 1640 complete medium
[0093] ELISA antibody:
[0094] Biotinylated2019 n-Cov Spike S-His (KACTUS company)
[0095] Spike S2(SARS-Cov-2)(HEK293)(Beijing Sino Biological Technology Co., Ltd.)
[0096] GoaAnti-Mouse IgG-HRP (1:10000)
[0097] Mice: female Balb / c mice (5 weeks, about 25g)
[0098] method:
[0099] Take DC-pCDH-K-S2-CD4, DC-PCDH-K-S2, DC-PCDH-K-S and other three kinds of cells each 1×10 8 cell, and then resuspend the cells with 1ml of PBS.
[0100] Animal experiments were divided into six groups: ①PBS group, ②DC-S2-CD4 group, ③DC-PCDH-K-S2 group, ④DC-PCDH-K-S group, 4 mice in each group, each mouse was injected with 2×10 7 cells.
[0101] After 14 days, take 200 μl of blood from each mouse to separate serum: place the collected serum in a 37°C incubator for 1 hour, and then centrifuge at 3000 rpm for 20 minutes. After the centrifugation, transfer the upper serum to a labeled 1.5ml centrifuge tube,...
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