CN116970624A - 抗SARS-CoV-2及变异株感染的腺病毒疫苗 - Google Patents
抗SARS-CoV-2及变异株感染的腺病毒疫苗 Download PDFInfo
- Publication number
- CN116970624A CN116970624A CN202310726118.1A CN202310726118A CN116970624A CN 116970624 A CN116970624 A CN 116970624A CN 202310726118 A CN202310726118 A CN 202310726118A CN 116970624 A CN116970624 A CN 116970624A
- Authority
- CN
- China
- Prior art keywords
- seq
- protein
- cov
- polynucleotide
- vaccine
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Pending
Links
- 241001678559 COVID-19 virus Species 0.000 title claims abstract description 41
- 208000015181 infectious disease Diseases 0.000 title claims abstract description 13
- 229940021704 adenovirus vaccine Drugs 0.000 title abstract description 29
- 101710139375 Corneodesmosin Proteins 0.000 claims abstract description 55
- 229960005486 vaccine Drugs 0.000 claims abstract description 49
- 241000701161 unidentified adenovirus Species 0.000 claims abstract description 35
- 241000711573 Coronaviridae Species 0.000 claims abstract description 15
- 230000002950 deficient Effects 0.000 claims abstract description 14
- 230000010076 replication Effects 0.000 claims abstract description 14
- 230000002265 prevention Effects 0.000 claims abstract description 5
- 239000003814 drug Substances 0.000 claims abstract description 4
- 102100031673 Corneodesmosin Human genes 0.000 claims description 45
- 108091033319 polynucleotide Proteins 0.000 claims description 39
- 239000002157 polynucleotide Substances 0.000 claims description 39
- 102000040430 polynucleotide Human genes 0.000 claims description 39
- 239000002773 nucleotide Substances 0.000 claims description 22
- 125000003729 nucleotide group Chemical group 0.000 claims description 22
- 108010076504 Protein Sorting Signals Proteins 0.000 claims description 21
- 150000001413 amino acids Chemical class 0.000 claims description 18
- 238000000034 method Methods 0.000 claims description 17
- 230000035772 mutation Effects 0.000 claims description 14
- 239000013598 vector Substances 0.000 claims description 13
- 239000013612 plasmid Substances 0.000 claims description 12
- 239000013600 plasmid vector Substances 0.000 claims description 8
- 239000007923 nasal drop Substances 0.000 claims description 6
- 241000282577 Pan troglodytes Species 0.000 claims description 5
- -1 carrier Substances 0.000 claims description 5
- 241001135569 Human adenovirus 5 Species 0.000 claims description 4
- 201000003176 Severe Acute Respiratory Syndrome Diseases 0.000 claims description 4
- 230000003321 amplification Effects 0.000 claims description 4
- 239000007922 nasal spray Substances 0.000 claims description 4
- 238000003199 nucleic acid amplification method Methods 0.000 claims description 4
- 238000000746 purification Methods 0.000 claims description 4
- 108090001126 Furin Proteins 0.000 claims description 3
- 102100034013 Gamma-glutamyl phosphate reductase Human genes 0.000 claims description 3
- 101001133924 Homo sapiens Gamma-glutamyl phosphate reductase Proteins 0.000 claims description 3
- 101000801481 Homo sapiens Tissue-type plasminogen activator Proteins 0.000 claims description 3
- 108091028043 Nucleic acid sequence Proteins 0.000 claims description 3
- 206010062106 Respiratory tract infection viral Diseases 0.000 claims description 3
- 239000002671 adjuvant Substances 0.000 claims description 3
- 238000003776 cleavage reaction Methods 0.000 claims description 3
- 238000012258 culturing Methods 0.000 claims description 3
- 238000010255 intramuscular injection Methods 0.000 claims description 3
- 239000007927 intramuscular injection Substances 0.000 claims description 3
- 229940100662 nasal drops Drugs 0.000 claims description 3
- 230000007017 scission Effects 0.000 claims description 3
- 230000004071 biological effect Effects 0.000 claims description 2
- 239000003085 diluting agent Substances 0.000 claims description 2
- 230000001717 pathogenic effect Effects 0.000 claims description 2
- 239000000546 pharmaceutical excipient Substances 0.000 claims description 2
- 239000007921 spray Substances 0.000 claims description 2
- 125000003275 alpha amino acid group Chemical group 0.000 claims 4
- 239000003795 chemical substances by application Substances 0.000 claims 1
- 238000004519 manufacturing process Methods 0.000 claims 1
- 241000700605 Viruses Species 0.000 abstract description 37
- 208000025721 COVID-19 Diseases 0.000 abstract description 2
- 208000001528 Coronaviridae Infections Diseases 0.000 abstract description 2
- 208000037847 SARS-CoV-2-infection Diseases 0.000 abstract description 2
- 230000000694 effects Effects 0.000 abstract description 2
- 210000004027 cell Anatomy 0.000 description 53
- 101710137302 Surface antigen S Proteins 0.000 description 17
- 238000001514 detection method Methods 0.000 description 16
- 108090000623 proteins and genes Proteins 0.000 description 16
- 238000012360 testing method Methods 0.000 description 14
- 210000001519 tissue Anatomy 0.000 description 11
- AIYUHDOJVYHVIT-UHFFFAOYSA-M caesium chloride Chemical compound [Cl-].[Cs+] AIYUHDOJVYHVIT-UHFFFAOYSA-M 0.000 description 10
- 239000006228 supernatant Substances 0.000 description 10
- 210000001744 T-lymphocyte Anatomy 0.000 description 9
- 210000002966 serum Anatomy 0.000 description 9
- 230000003472 neutralizing effect Effects 0.000 description 8
- 102000004169 proteins and genes Human genes 0.000 description 8
- 239000000243 solution Substances 0.000 description 8
- 239000012530 fluid Substances 0.000 description 7
- 210000004072 lung Anatomy 0.000 description 7
- 241000699670 Mus sp. Species 0.000 description 6
- ONIBWKKTOPOVIA-UHFFFAOYSA-N Proline Natural products OC(=O)C1CCCN1 ONIBWKKTOPOVIA-UHFFFAOYSA-N 0.000 description 6
- 229940096437 Protein S Drugs 0.000 description 6
- 241001112090 Pseudovirus Species 0.000 description 6
- FAPWRFPIFSIZLT-UHFFFAOYSA-M Sodium chloride Chemical compound [Na+].[Cl-] FAPWRFPIFSIZLT-UHFFFAOYSA-M 0.000 description 6
- 101710198474 Spike protein Proteins 0.000 description 6
- 239000002609 medium Substances 0.000 description 6
- 108091003079 Bovine Serum Albumin Proteins 0.000 description 5
- 239000000427 antigen Substances 0.000 description 5
- 108091007433 antigens Proteins 0.000 description 5
- 102000036639 antigens Human genes 0.000 description 5
- 210000004369 blood Anatomy 0.000 description 5
- 239000008280 blood Substances 0.000 description 5
- 238000004113 cell culture Methods 0.000 description 5
- 238000000684 flow cytometry Methods 0.000 description 5
- 230000003053 immunization Effects 0.000 description 5
- 238000002649 immunization Methods 0.000 description 5
- 230000008569 process Effects 0.000 description 5
- 108020004705 Codon Proteins 0.000 description 4
- 241000196324 Embryophyta Species 0.000 description 4
- 108010001336 Horseradish Peroxidase Proteins 0.000 description 4
- 229920001213 Polysorbate 20 Polymers 0.000 description 4
- 229940098773 bovine serum albumin Drugs 0.000 description 4
- 239000000872 buffer Substances 0.000 description 4
- HVYWMOMLDIMFJA-DPAQBDIFSA-N cholesterol Chemical compound C1C=C2C[C@@H](O)CC[C@]2(C)[C@@H]2[C@@H]1[C@@H]1CC[C@H]([C@H](C)CCCC(C)C)[C@@]1(C)CC2 HVYWMOMLDIMFJA-DPAQBDIFSA-N 0.000 description 4
- 238000001914 filtration Methods 0.000 description 4
- 238000004255 ion exchange chromatography Methods 0.000 description 4
- 239000000256 polyoxyethylene sorbitan monolaurate Substances 0.000 description 4
- 235000010486 polyoxyethylene sorbitan monolaurate Nutrition 0.000 description 4
- 238000002360 preparation method Methods 0.000 description 4
- 239000000047 product Substances 0.000 description 4
- QKNYBSVHEMOAJP-UHFFFAOYSA-N 2-amino-2-(hydroxymethyl)propane-1,3-diol;hydron;chloride Chemical compound Cl.OCC(N)(CO)CO QKNYBSVHEMOAJP-UHFFFAOYSA-N 0.000 description 3
- 239000006144 Dulbecco’s modified Eagle's medium Substances 0.000 description 3
- PEDCQBHIVMGVHV-UHFFFAOYSA-N Glycerine Chemical compound OCC(O)CO PEDCQBHIVMGVHV-UHFFFAOYSA-N 0.000 description 3
- KZSNJWFQEVHDMF-BYPYZUCNSA-N L-valine Chemical compound CC(C)[C@H](N)C(O)=O KZSNJWFQEVHDMF-BYPYZUCNSA-N 0.000 description 3
- KDXKERNSBIXSRK-UHFFFAOYSA-N Lysine Natural products NCCCCC(N)C(O)=O KDXKERNSBIXSRK-UHFFFAOYSA-N 0.000 description 3
- 239000004472 Lysine Substances 0.000 description 3
- 101150010882 S gene Proteins 0.000 description 3
- KZSNJWFQEVHDMF-UHFFFAOYSA-N Valine Natural products CC(C)C(N)C(O)=O KZSNJWFQEVHDMF-UHFFFAOYSA-N 0.000 description 3
- ZMJPCIAEJKVKMQ-UHFFFAOYSA-M [4-[[4-[benzyl(methyl)amino]phenyl]-[4-(dimethylamino)phenyl]methylidene]cyclohexa-2,5-dien-1-ylidene]-dimethylazanium;chloride Chemical compound [Cl-].C1=CC(N(C)C)=CC=C1C(C=1C=CC(=CC=1)N(C)CC=1C=CC=CC=1)=C1C=CC(=[N+](C)C)C=C1 ZMJPCIAEJKVKMQ-UHFFFAOYSA-M 0.000 description 3
- 238000003556 assay Methods 0.000 description 3
- 230000015572 biosynthetic process Effects 0.000 description 3
- 239000007853 buffer solution Substances 0.000 description 3
- 125000002091 cationic group Chemical group 0.000 description 3
- 239000006285 cell suspension Substances 0.000 description 3
- 238000006243 chemical reaction Methods 0.000 description 3
- 239000003153 chemical reaction reagent Substances 0.000 description 3
- 239000012228 culture supernatant Substances 0.000 description 3
- 238000010790 dilution Methods 0.000 description 3
- 239000012895 dilution Substances 0.000 description 3
- 238000010828 elution Methods 0.000 description 3
- 230000003203 everyday effect Effects 0.000 description 3
- 239000012091 fetal bovine serum Substances 0.000 description 3
- 150000004676 glycans Chemical class 0.000 description 3
- 239000012528 membrane Substances 0.000 description 3
- 238000005457 optimization Methods 0.000 description 3
- 239000008188 pellet Substances 0.000 description 3
- 238000003752 polymerase chain reaction Methods 0.000 description 3
- 229920001282 polysaccharide Polymers 0.000 description 3
- 239000005017 polysaccharide Substances 0.000 description 3
- 230000001681 protective effect Effects 0.000 description 3
- 239000011780 sodium chloride Substances 0.000 description 3
- 230000009870 specific binding Effects 0.000 description 3
- 238000010186 staining Methods 0.000 description 3
- 238000003786 synthesis reaction Methods 0.000 description 3
- 238000005199 ultracentrifugation Methods 0.000 description 3
- 239000004474 valine Substances 0.000 description 3
- 238000005406 washing Methods 0.000 description 3
- MZOFCQQQCNRIBI-VMXHOPILSA-N (3s)-4-[[(2s)-1-[[(2s)-1-[[(1s)-1-carboxy-2-hydroxyethyl]amino]-4-methyl-1-oxopentan-2-yl]amino]-5-(diaminomethylideneamino)-1-oxopentan-2-yl]amino]-3-[[2-[[(2s)-2,6-diaminohexanoyl]amino]acetyl]amino]-4-oxobutanoic acid Chemical compound OC[C@@H](C(O)=O)NC(=O)[C@H](CC(C)C)NC(=O)[C@H](CCCN=C(N)N)NC(=O)[C@H](CC(O)=O)NC(=O)CNC(=O)[C@@H](N)CCCCN MZOFCQQQCNRIBI-VMXHOPILSA-N 0.000 description 2
- 238000011725 BALB/c mouse Methods 0.000 description 2
- 102100032912 CD44 antigen Human genes 0.000 description 2
- 108090000695 Cytokines Proteins 0.000 description 2
- 102000004127 Cytokines Human genes 0.000 description 2
- 241001533413 Deltavirus Species 0.000 description 2
- 238000002965 ELISA Methods 0.000 description 2
- 102000004961 Furin Human genes 0.000 description 2
- WQZGKKKJIJFFOK-GASJEMHNSA-N Glucose Natural products OC[C@H]1OC(O)[C@H](O)[C@@H](O)[C@@H]1O WQZGKKKJIJFFOK-GASJEMHNSA-N 0.000 description 2
- 101000868273 Homo sapiens CD44 antigen Proteins 0.000 description 2
- 102100037850 Interferon gamma Human genes 0.000 description 2
- 108010074328 Interferon-gamma Proteins 0.000 description 2
- 239000005089 Luciferase Substances 0.000 description 2
- 239000012124 Opti-MEM Substances 0.000 description 2
- 238000012408 PCR amplification Methods 0.000 description 2
- LCTONWCANYUPML-UHFFFAOYSA-N Pyruvic acid Chemical compound CC(=O)C(O)=O LCTONWCANYUPML-UHFFFAOYSA-N 0.000 description 2
- 108091005634 SARS-CoV-2 receptor-binding domains Proteins 0.000 description 2
- 229920002684 Sepharose Polymers 0.000 description 2
- 108060008682 Tumor Necrosis Factor Proteins 0.000 description 2
- 102000000852 Tumor Necrosis Factor-alpha Human genes 0.000 description 2
- 238000002835 absorbance Methods 0.000 description 2
- AZDRQVAHHNSJOQ-UHFFFAOYSA-N alumane Chemical class [AlH3] AZDRQVAHHNSJOQ-UHFFFAOYSA-N 0.000 description 2
- 230000009286 beneficial effect Effects 0.000 description 2
- 159000000007 calcium salts Chemical class 0.000 description 2
- 235000012000 cholesterol Nutrition 0.000 description 2
- 238000012217 deletion Methods 0.000 description 2
- 230000037430 deletion Effects 0.000 description 2
- 238000004925 denaturation Methods 0.000 description 2
- 230000036425 denaturation Effects 0.000 description 2
- 238000010586 diagram Methods 0.000 description 2
- 239000000945 filler Substances 0.000 description 2
- MHMNJMPURVTYEJ-UHFFFAOYSA-N fluorescein-5-isothiocyanate Chemical compound O1C(=O)C2=CC(N=C=S)=CC=C2C21C1=CC=C(O)C=C1OC1=CC(O)=CC=C21 MHMNJMPURVTYEJ-UHFFFAOYSA-N 0.000 description 2
- 210000001280 germinal center Anatomy 0.000 description 2
- 239000008103 glucose Substances 0.000 description 2
- 230000036039 immunity Effects 0.000 description 2
- 239000000568 immunological adjuvant Substances 0.000 description 2
- 230000003834 intracellular effect Effects 0.000 description 2
- 150000002632 lipids Chemical class 0.000 description 2
- 239000002502 liposome Substances 0.000 description 2
- 238000011068 loading method Methods 0.000 description 2
- 239000000463 material Substances 0.000 description 2
- 210000005015 mediastinal lymph node Anatomy 0.000 description 2
- 238000002156 mixing Methods 0.000 description 2
- 238000004806 packaging method and process Methods 0.000 description 2
- 239000002245 particle Substances 0.000 description 2
- 125000001500 prolyl group Chemical group [H]N1C([H])(C(=O)[*])C([H])([H])C([H])([H])C1([H])[H] 0.000 description 2
- 210000002345 respiratory system Anatomy 0.000 description 2
- 230000004044 response Effects 0.000 description 2
- 238000005070 sampling Methods 0.000 description 2
- 229930182490 saponin Natural products 0.000 description 2
- 150000007949 saponins Chemical class 0.000 description 2
- 235000017709 saponins Nutrition 0.000 description 2
- UCSJYZPVAKXKNQ-HZYVHMACSA-N streptomycin Chemical compound CN[C@H]1[C@H](O)[C@@H](O)[C@H](CO)O[C@H]1O[C@@H]1[C@](C=O)(O)[C@H](C)O[C@H]1O[C@@H]1[C@@H](NC(N)=N)[C@H](O)[C@@H](NC(N)=N)[C@H](O)[C@H]1O UCSJYZPVAKXKNQ-HZYVHMACSA-N 0.000 description 2
- 238000010257 thawing Methods 0.000 description 2
- 238000002255 vaccination Methods 0.000 description 2
- 230000009385 viral infection Effects 0.000 description 2
- 230000003612 virological effect Effects 0.000 description 2
- HBZBAMXERPYTFS-SECBINFHSA-N (4S)-2-(6,7-dihydro-5H-pyrrolo[3,2-f][1,3]benzothiazol-2-yl)-4,5-dihydro-1,3-thiazole-4-carboxylic acid Chemical compound OC(=O)[C@H]1CSC(=N1)c1nc2cc3CCNc3cc2s1 HBZBAMXERPYTFS-SECBINFHSA-N 0.000 description 1
- YYGNTYWPHWGJRM-UHFFFAOYSA-N (6E,10E,14E,18E)-2,6,10,15,19,23-hexamethyltetracosa-2,6,10,14,18,22-hexaene Chemical compound CC(C)=CCCC(C)=CCCC(C)=CCCC=C(C)CCC=C(C)CCC=C(C)C YYGNTYWPHWGJRM-UHFFFAOYSA-N 0.000 description 1
- LDGWQMRUWMSZIU-LQDDAWAPSA-M 2,3-bis[(z)-octadec-9-enoxy]propyl-trimethylazanium;chloride Chemical compound [Cl-].CCCCCCCC\C=C/CCCCCCCCOCC(C[N+](C)(C)C)OCCCCCCCC\C=C/CCCCCCCC LDGWQMRUWMSZIU-LQDDAWAPSA-M 0.000 description 1
- 108010042708 Acetylmuramyl-Alanyl-Isoglutamine Proteins 0.000 description 1
- 102000053723 Angiotensin-converting enzyme 2 Human genes 0.000 description 1
- 108090000975 Angiotensin-converting enzyme 2 Proteins 0.000 description 1
- 241001061264 Astragalus Species 0.000 description 1
- 102100024222 B-lymphocyte antigen CD19 Human genes 0.000 description 1
- 102100031658 C-X-C chemokine receptor type 5 Human genes 0.000 description 1
- CXRFDZFCGOPDTD-UHFFFAOYSA-M Cetrimide Chemical compound [Br-].CCCCCCCCCCCCCC[N+](C)(C)C CXRFDZFCGOPDTD-UHFFFAOYSA-M 0.000 description 1
- LZZYPRNAOMGNLH-UHFFFAOYSA-M Cetrimonium bromide Chemical compound [Br-].CCCCCCCCCCCCCCCC[N+](C)(C)C LZZYPRNAOMGNLH-UHFFFAOYSA-M 0.000 description 1
- VEXZGXHMUGYJMC-UHFFFAOYSA-M Chloride anion Chemical compound [Cl-] VEXZGXHMUGYJMC-UHFFFAOYSA-M 0.000 description 1
- 102000009016 Cholera Toxin Human genes 0.000 description 1
- 108010049048 Cholera Toxin Proteins 0.000 description 1
- 102000029816 Collagenase Human genes 0.000 description 1
- 108060005980 Collagenase Proteins 0.000 description 1
- 102100025137 Early activation antigen CD69 Human genes 0.000 description 1
- 108010067770 Endopeptidase K Proteins 0.000 description 1
- JZNWSCPGTDBMEW-UHFFFAOYSA-N Glycerophosphorylethanolamin Natural products NCCOP(O)(=O)OCC(O)CO JZNWSCPGTDBMEW-UHFFFAOYSA-N 0.000 description 1
- 101710114810 Glycoprotein Proteins 0.000 description 1
- 102000004457 Granulocyte-Macrophage Colony-Stimulating Factor Human genes 0.000 description 1
- 108010017213 Granulocyte-Macrophage Colony-Stimulating Factor Proteins 0.000 description 1
- 101000980825 Homo sapiens B-lymphocyte antigen CD19 Proteins 0.000 description 1
- 101000922405 Homo sapiens C-X-C chemokine receptor type 5 Proteins 0.000 description 1
- 101000934374 Homo sapiens Early activation antigen CD69 Proteins 0.000 description 1
- 101000611023 Homo sapiens Tumor necrosis factor receptor superfamily member 6 Proteins 0.000 description 1
- 108010002350 Interleukin-2 Proteins 0.000 description 1
- 108060001084 Luciferase Proteins 0.000 description 1
- 108010052285 Membrane Proteins Proteins 0.000 description 1
- 102000018697 Membrane Proteins Human genes 0.000 description 1
- 241001465754 Metazoa Species 0.000 description 1
- 241000127282 Middle East respiratory syndrome-related coronavirus Species 0.000 description 1
- 241000699666 Mus <mouse, genus> Species 0.000 description 1
- 101710163270 Nuclease Proteins 0.000 description 1
- 108090001074 Nucleocapsid Proteins Proteins 0.000 description 1
- 229910019142 PO4 Inorganic materials 0.000 description 1
- 229930182555 Penicillin Natural products 0.000 description 1
- JGSARLDLIJGVTE-MBNYWOFBSA-N Penicillin G Chemical compound N([C@H]1[C@H]2SC([C@@H](N2C1=O)C(O)=O)(C)C)C(=O)CC1=CC=CC=C1 JGSARLDLIJGVTE-MBNYWOFBSA-N 0.000 description 1
- 108010067902 Peptide Library Proteins 0.000 description 1
- 101000933967 Pseudomonas phage KPP25 Major capsid protein Proteins 0.000 description 1
- 208000035415 Reinfection Diseases 0.000 description 1
- 108700008625 Reporter Genes Proteins 0.000 description 1
- 241000315672 SARS coronavirus Species 0.000 description 1
- 101710167605 Spike glycoprotein Proteins 0.000 description 1
- 229930006000 Sucrose Natural products 0.000 description 1
- CZMRCDWAGMRECN-UGDNZRGBSA-N Sucrose Chemical compound O[C@H]1[C@H](O)[C@@H](CO)O[C@@]1(CO)O[C@@H]1[C@H](O)[C@@H](O)[C@H](O)[C@@H](CO)O1 CZMRCDWAGMRECN-UGDNZRGBSA-N 0.000 description 1
- 230000005867 T cell response Effects 0.000 description 1
- BHEOSNUKNHRBNM-UHFFFAOYSA-N Tetramethylsqualene Natural products CC(=C)C(C)CCC(=C)C(C)CCC(C)=CCCC=C(C)CCC(C)C(=C)CCC(C)C(C)=C BHEOSNUKNHRBNM-UHFFFAOYSA-N 0.000 description 1
- 239000007983 Tris buffer Substances 0.000 description 1
- 102100040403 Tumor necrosis factor receptor superfamily member 6 Human genes 0.000 description 1
- 208000036142 Viral infection Diseases 0.000 description 1
- 230000006978 adaptation Effects 0.000 description 1
- 229940037003 alum Drugs 0.000 description 1
- WNROFYMDJYEPJX-UHFFFAOYSA-K aluminium hydroxide Chemical compound [OH-].[OH-].[OH-].[Al+3] WNROFYMDJYEPJX-UHFFFAOYSA-K 0.000 description 1
- DIZPMCHEQGEION-UHFFFAOYSA-H aluminium sulfate (anhydrous) Chemical compound [Al+3].[Al+3].[O-]S([O-])(=O)=O.[O-]S([O-])(=O)=O.[O-]S([O-])(=O)=O DIZPMCHEQGEION-UHFFFAOYSA-H 0.000 description 1
- 238000005571 anion exchange chromatography Methods 0.000 description 1
- 238000000137 annealing Methods 0.000 description 1
- 239000003242 anti bacterial agent Substances 0.000 description 1
- 229940088710 antibiotic agent Drugs 0.000 description 1
- 235000006533 astragalus Nutrition 0.000 description 1
- 239000001506 calcium phosphate Substances 0.000 description 1
- 239000002775 capsule Substances 0.000 description 1
- 239000000969 carrier Substances 0.000 description 1
- 230000001413 cellular effect Effects 0.000 description 1
- 230000007969 cellular immunity Effects 0.000 description 1
- 238000005119 centrifugation Methods 0.000 description 1
- 238000010367 cloning Methods 0.000 description 1
- 229960002424 collagenase Drugs 0.000 description 1
- 239000012141 concentrate Substances 0.000 description 1
- 238000010276 construction Methods 0.000 description 1
- 238000009295 crossflow filtration Methods 0.000 description 1
- 238000005520 cutting process Methods 0.000 description 1
- 230000009089 cytolysis Effects 0.000 description 1
- 230000034994 death Effects 0.000 description 1
- 239000003599 detergent Substances 0.000 description 1
- 238000011161 development Methods 0.000 description 1
- 238000000502 dialysis Methods 0.000 description 1
- PSLWZOIUBRXAQW-UHFFFAOYSA-M dimethyl(dioctadecyl)azanium;bromide Chemical compound [Br-].CCCCCCCCCCCCCCCCCC[N+](C)(C)CCCCCCCCCCCCCCCCCC PSLWZOIUBRXAQW-UHFFFAOYSA-M 0.000 description 1
- MWRBNPKJOOWZPW-CLFAGFIQSA-N dioleoyl phosphatidylethanolamine Chemical compound CCCCCCCC\C=C/CCCCCCCC(=O)OCC(COP(O)(=O)OCCN)OC(=O)CCCCCCC\C=C/CCCCCCCC MWRBNPKJOOWZPW-CLFAGFIQSA-N 0.000 description 1
- 238000006073 displacement reaction Methods 0.000 description 1
- PRAKJMSDJKAYCZ-UHFFFAOYSA-N dodecahydrosqualene Natural products CC(C)CCCC(C)CCCC(C)CCCCC(C)CCCC(C)CCCC(C)C PRAKJMSDJKAYCZ-UHFFFAOYSA-N 0.000 description 1
- XJWSAJYUBXQQDR-UHFFFAOYSA-M dodecyltrimethylammonium bromide Chemical compound [Br-].CCCCCCCCCCCC[N+](C)(C)C XJWSAJYUBXQQDR-UHFFFAOYSA-M 0.000 description 1
- 239000002552 dosage form Substances 0.000 description 1
- 238000001962 electrophoresis Methods 0.000 description 1
- 230000017188 evasion or tolerance of host immune response Effects 0.000 description 1
- 239000001963 growth medium Substances 0.000 description 1
- 230000036541 health Effects 0.000 description 1
- 239000012145 high-salt buffer Substances 0.000 description 1
- 230000028993 immune response Effects 0.000 description 1
- 230000001571 immunoadjuvant effect Effects 0.000 description 1
- 238000011081 inoculation Methods 0.000 description 1
- 239000007788 liquid Substances 0.000 description 1
- 230000005923 long-lasting effect Effects 0.000 description 1
- 210000004698 lymphocyte Anatomy 0.000 description 1
- 239000006166 lysate Substances 0.000 description 1
- 125000003588 lysine group Chemical class [H]N([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])(N([H])[H])C(*)=O 0.000 description 1
- 230000002934 lysing effect Effects 0.000 description 1
- 210000001370 mediastinum Anatomy 0.000 description 1
- 238000012986 modification Methods 0.000 description 1
- 230000004048 modification Effects 0.000 description 1
- 229940035032 monophosphoryl lipid a Drugs 0.000 description 1
- BSOQXXWZTUDTEL-ZUYCGGNHSA-N muramyl dipeptide Chemical compound OC(=O)CC[C@H](C(N)=O)NC(=O)[C@H](C)NC(=O)[C@@H](C)O[C@H]1[C@H](O)[C@@H](CO)O[C@@H](O)[C@@H]1NC(C)=O BSOQXXWZTUDTEL-ZUYCGGNHSA-N 0.000 description 1
- 125000001446 muramyl group Chemical group N[C@@H](C=O)[C@@H](O[C@@H](C(=O)*)C)[C@H](O)[C@H](O)CO 0.000 description 1
- 229940100661 nasal inhalant Drugs 0.000 description 1
- 229940097496 nasal spray Drugs 0.000 description 1
- 238000006386 neutralization reaction Methods 0.000 description 1
- 239000007764 o/w emulsion Substances 0.000 description 1
- 230000008520 organization Effects 0.000 description 1
- 244000052769 pathogen Species 0.000 description 1
- 229940049954 penicillin Drugs 0.000 description 1
- 230000002085 persistent effect Effects 0.000 description 1
- WTJKGGKOPKCXLL-RRHRGVEJSA-N phosphatidylcholine Chemical compound CCCCCCCCCCCCCCCC(=O)OC[C@H](COP([O-])(=O)OCC[N+](C)(C)C)OC(=O)CCCCCCCC=CCCCCCCCC WTJKGGKOPKCXLL-RRHRGVEJSA-N 0.000 description 1
- 150000008104 phosphatidylethanolamines Chemical class 0.000 description 1
- 235000010482 polyoxyethylene sorbitan monooleate Nutrition 0.000 description 1
- 229920000053 polysorbate 80 Polymers 0.000 description 1
- 239000002244 precipitate Substances 0.000 description 1
- 125000001436 propyl group Chemical group [H]C([*])([H])C([H])([H])C([H])([H])[H] 0.000 description 1
- 229940023143 protein vaccine Drugs 0.000 description 1
- 229940107700 pyruvic acid Drugs 0.000 description 1
- 239000001397 quillaja saponaria molina bark Substances 0.000 description 1
- 238000005215 recombination Methods 0.000 description 1
- 230000006798 recombination Effects 0.000 description 1
- 238000011084 recovery Methods 0.000 description 1
- 230000003248 secreting effect Effects 0.000 description 1
- 230000028327 secretion Effects 0.000 description 1
- 238000012163 sequencing technique Methods 0.000 description 1
- 238000004513 sizing Methods 0.000 description 1
- 229940031439 squalene Drugs 0.000 description 1
- TUHBEKDERLKLEC-UHFFFAOYSA-N squalene Natural products CC(=CCCC(=CCCC(=CCCC=C(/C)CCC=C(/C)CC=C(C)C)C)C)C TUHBEKDERLKLEC-UHFFFAOYSA-N 0.000 description 1
- 230000001954 sterilising effect Effects 0.000 description 1
- 229960005322 streptomycin Drugs 0.000 description 1
- 239000000126 substance Substances 0.000 description 1
- 238000006467 substitution reaction Methods 0.000 description 1
- 239000005720 sucrose Substances 0.000 description 1
- 239000000725 suspension Substances 0.000 description 1
- 210000004233 talus Anatomy 0.000 description 1
- 231100000331 toxic Toxicity 0.000 description 1
- 230000002588 toxic effect Effects 0.000 description 1
- 239000003053 toxin Substances 0.000 description 1
- 231100000765 toxin Toxicity 0.000 description 1
- 238000001890 transfection Methods 0.000 description 1
- QORWJWZARLRLPR-UHFFFAOYSA-H tricalcium bis(phosphate) Chemical group [Ca+2].[Ca+2].[Ca+2].[O-]P([O-])([O-])=O.[O-]P([O-])([O-])=O QORWJWZARLRLPR-UHFFFAOYSA-H 0.000 description 1
- 229940078499 tricalcium phosphate Drugs 0.000 description 1
- 229910000391 tricalcium phosphate Inorganic materials 0.000 description 1
- 235000019731 tricalcium phosphate Nutrition 0.000 description 1
- LENZDBCJOHFCAS-UHFFFAOYSA-N tris Chemical compound OCC(N)(CO)CO LENZDBCJOHFCAS-UHFFFAOYSA-N 0.000 description 1
- 238000000108 ultra-filtration Methods 0.000 description 1
- 229940125575 vaccine candidate Drugs 0.000 description 1
- 210000003462 vein Anatomy 0.000 description 1
- 230000001018 virulence Effects 0.000 description 1
- 239000011534 wash buffer Substances 0.000 description 1
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 description 1
- 238000001262 western blot Methods 0.000 description 1
Classifications
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07K—PEPTIDES
- C07K14/00—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
- C07K14/005—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from viruses
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K39/00—Medicinal preparations containing antigens or antibodies
- A61K39/12—Viral antigens
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K39/00—Medicinal preparations containing antigens or antibodies
- A61K39/39—Medicinal preparations containing antigens or antibodies characterised by the immunostimulating additives, e.g. chemical adjuvants
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- A61P31/00—Antiinfectives, i.e. antibiotics, antiseptics, chemotherapeutics
- A61P31/12—Antivirals
- A61P31/14—Antivirals for RNA viruses
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N15/00—Mutation or genetic engineering; DNA or RNA concerning genetic engineering, vectors, e.g. plasmids, or their isolation, preparation or purification; Use of hosts therefor
- C12N15/09—Recombinant DNA-technology
- C12N15/63—Introduction of foreign genetic material using vectors; Vectors; Use of hosts therefor; Regulation of expression
- C12N15/79—Vectors or expression systems specially adapted for eukaryotic hosts
- C12N15/85—Vectors or expression systems specially adapted for eukaryotic hosts for animal cells
- C12N15/86—Viral vectors
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K39/00—Medicinal preparations containing antigens or antibodies
- A61K2039/51—Medicinal preparations containing antigens or antibodies comprising whole cells, viruses or DNA/RNA
- A61K2039/525—Virus
- A61K2039/5256—Virus expressing foreign proteins
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K39/00—Medicinal preparations containing antigens or antibodies
- A61K2039/54—Medicinal preparations containing antigens or antibodies characterised by the route of administration
- A61K2039/541—Mucosal route
- A61K2039/543—Mucosal route intranasal
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N2710/00—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA dsDNA viruses
- C12N2710/00011—Details
- C12N2710/10011—Adenoviridae
- C12N2710/10311—Mastadenovirus, e.g. human or simian adenoviruses
- C12N2710/10334—Use of virus or viral component as vaccine, e.g. live-attenuated or inactivated virus, VLP, viral protein
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N2710/00—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA dsDNA viruses
- C12N2710/00011—Details
- C12N2710/10011—Adenoviridae
- C12N2710/10311—Mastadenovirus, e.g. human or simian adenoviruses
- C12N2710/10341—Use of virus, viral particle or viral elements as a vector
- C12N2710/10343—Use of virus, viral particle or viral elements as a vector viral genome or elements thereof as genetic vector
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N2770/00—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA ssRNA viruses positive-sense
- C12N2770/00011—Details
- C12N2770/20011—Coronaviridae
- C12N2770/20022—New viral proteins or individual genes, new structural or functional aspects of known viral proteins or genes
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N2770/00—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA ssRNA viruses positive-sense
- C12N2770/00011—Details
- C12N2770/20011—Coronaviridae
- C12N2770/20034—Use of virus or viral component as vaccine, e.g. live-attenuated or inactivated virus, VLP, viral protein
Landscapes
- Health & Medical Sciences (AREA)
- Life Sciences & Earth Sciences (AREA)
- Chemical & Material Sciences (AREA)
- Genetics & Genomics (AREA)
- Virology (AREA)
- Organic Chemistry (AREA)
- General Health & Medical Sciences (AREA)
- Medicinal Chemistry (AREA)
- Animal Behavior & Ethology (AREA)
- Microbiology (AREA)
- Molecular Biology (AREA)
- Veterinary Medicine (AREA)
- Public Health (AREA)
- Pharmacology & Pharmacy (AREA)
- Engineering & Computer Science (AREA)
- Wood Science & Technology (AREA)
- Epidemiology (AREA)
- General Engineering & Computer Science (AREA)
- General Chemical & Material Sciences (AREA)
- Bioinformatics & Cheminformatics (AREA)
- Biomedical Technology (AREA)
- Biophysics (AREA)
- Zoology (AREA)
- Immunology (AREA)
- Mycology (AREA)
- Chemical Kinetics & Catalysis (AREA)
- Biotechnology (AREA)
- Biochemistry (AREA)
- Gastroenterology & Hepatology (AREA)
- Proteomics, Peptides & Aminoacids (AREA)
- Physics & Mathematics (AREA)
- Plant Pathology (AREA)
- Communicable Diseases (AREA)
- Oncology (AREA)
- Nuclear Medicine, Radiotherapy & Molecular Imaging (AREA)
- Medicines Containing Antibodies Or Antigens For Use As Internal Diagnostic Agents (AREA)
Abstract
本发明涉及抗SARS‑CoV‑2及变异株感染的疫苗，特别包含一种广谱的抗SARS‑CoV‑2及变异株感染的腺病毒疫苗，属于医药领域。为解决针对SARS‑CoV‑2及其变异株感染尚缺乏有效预防和治疗药物的问题，本发明提供了以人5型复制缺陷腺病毒为载体的新型冠状病毒疫苗，特别包含一种广谱的抗SARS‑CoV‑2感染的疫苗，该疫苗以E1/E3缺失的复制缺陷型人5型腺病毒为载体，携带经过优化设计的SARS‑CoV‑2及其变异株S蛋白基因，从而帮助宿主抵抗冠状病毒感染，特别是对突变病毒有较好的防治效果。
Description
技术领域
本发明属于生物医药技术领域，具体涉及抗SARS-CoV-2及变异株感染的腺病毒疫苗。
背景技术
新型冠状病毒(SARS-Cov-2)是由世界卫生组织命名的一种新型的β属的冠状病毒。该病毒有包膜，颗粒呈圆形或椭圆形，常为多形性，直径60-140nm。其基因特征与SARS-CoV和MERS-CoV有明显区别，是一种以前尚未在人类中发现的新冠状病毒分支。目前，变异新冠病毒主要有五大种类：Alpha、Beta、Gamma、Delta、Omicron，Omicron变异株又分为BA.1、BA.2、BA.2.12.1、BA4和BA5等几种亚系。截止目前为止，新型冠状病毒SARS-CoV-2已导致全球5.45亿人感染，其中死亡人数已超630万人。
SARS-CoV-2的主要结构蛋白包括刺突蛋白(Spike，S)、信使蛋白(Envelop，E)、膜蛋白(Membrane，M)和核衣壳蛋白(Nucleocapsid，N)，其中，S蛋白在病毒的感染和毒力中起着关键作用，常被用来作为疫苗的抗原。由于SARS-CoV-2变异株含有多个突变位点，病毒的S蛋白也含有多个突变位点，导致变异株一定程度上能够逃逸新冠原型株(即“新冠病毒野生株”)疫苗激发的抗体，导致新冠疫苗失效或保护力降低，为疫情防控工作带来巨大压力。因此，开发针对各变异的SARS-CoV-2病毒的疫苗，特别对各变异的SARS-CoV-2病毒的广谱疫苗，对于新型冠状病毒的防治非常重要。
发明内容
本发明旨在至少解决现有技术中存在的技术问题之一。为此，本发明的目的在于提供抗SARS-CoV-2及变异株感染的腺病毒载体。本发明的另一目的在于提供利用所述腺病毒载体制备的用于预防和/或治疗SARS-CoV-2或其变异株感染的疫苗。
本发明首先提供了编码SARS-CoV-2或变异株S蛋白基因的多核苷酸，所述的多核苷酸序列如SEQ ID No.3所示；或者在SEQ ID No.3核苷酸序列经取代和/或缺失和/或插入至少一个核苷酸获得的，且与SEQ ID No.3具有相同或相似的生物学活性的核苷酸序列。
进一步地，所述多核苷酸序列包括S蛋白或其突变体自身的信号肽，所述自身信号肽核苷酸序列如SEQ ID No.1所示。
优选地，所述多核苷酸序列包括S蛋白或其突变体自身的信号肽和在自身信号肽前额外添加的人tPA信号肽。
其中，所述S蛋白或其突变体自身的信号肽可被其他信号肽替代。
进一步地，所述的多核苷酸序列编码的SARS-CoV-2变异株S蛋白是将编码SARS-CoV-2的S蛋白氨基酸中的第986位K替换为P，将987位V替换为P；所述编码SARS-CoV-2的S蛋白氨基酸序列如SEQ ID No.4所示。
在一个优选实施方案中，所述的多核苷酸序列编码的SARS-CoV-2变异株S蛋白还包括对Furin蛋白酶切位点RRAR突变成GSAS。
进一步地，所述多核苷酸序列选自新冠病毒野生型、Alpha、Beta、Gamma、Delta、Omicron毒株中的S蛋白核苷酸序列中的至少一种。
进一步地，所述多核苷酸序列选自SEQ ID No.3、SEQ ID No.5、SEQ ID No.7、SEQIDNo.9、SEQ ID No.11、SEQ ID No.13、SEQ ID No.15或SEQ ID No.17中的至少一种。
进一步地，上述多核苷酸序列编码的SARS-CoV-2及变异株S蛋白的氨基酸序列选自SEQ ID No.4、SEQ ID No.6、SEQ ID No.8、SEQ ID No.10、SEQ ID No.12、SEQ ID No.14、SEQ IDNo.16或SEQ ID No.18中的至少一种。
优选地，所述氨基酸序列是在Beta变异株的S蛋白突变位点的基础上加上Delta变异株S蛋白RBD的突变位点。
更优选地，所述多核苷酸序列选自SEQ ID No.9、SEQ ID No.11、SEQ ID No.13、SEQ IDNo.15或SEQ ID No.17中的至少一种。
更优选地，所述多核苷酸序列编码的SARS-CoV-2变异株S蛋白的氨基酸序列选自SEQ ID No.10、SEQ ID No.12、SEQ ID No.14、SEQ ID No.16或SEQ ID No.18中的至少一种。
本发明还提供能够表达上述多核苷酸的复制缺陷型重组腺病毒载体。
优选地，所述人复制缺陷型重组腺病毒载体选自人5型、35型、26型或/和黑猩猩AdC68型、AdC7型的复制缺陷型腺病毒，但不限于以上类型的腺病毒病毒载体。更优选地，选自E1、E3联合缺失的人5型复制缺陷型腺病毒。
本发明还提供预防和/或治疗SARS-CoV-2或其变异株感染的疫苗，其包括能够表达上述多核苷酸的人复制缺陷型重组腺病毒载体。
所述疫苗还包括药学上可接受的佐剂、载体、稀释剂或赋形剂。
进一步地，所述的佐剂为免疫佐剂。
优选地，所述的免疫佐剂选自如下至少一种：铝盐、钙盐、植物皂苷、植物多糖、单磷酸脂质A、胞壁酰二肽、胞壁酰三肽、角鲨烯水包油乳剂、重组霍乱毒素、GM-CSF细胞因子、脂质、阳离子脂质体材料、CpG ODN。
进一步地，所述的铝盐选自氢氧化铝、明矾中至少一种。
进一步地，所述的钙盐为磷酸三钙。
进一步地，所述的植物皂苷为QS–21或ISCOM。
进一步地，所述的植物多糖为黄芪多糖。
进一步地，所述的脂质选自如下至少一种：磷脂酰乙醇胺、磷脂酰胆碱、胆固醇、二油酰基磷脂酰乙醇胺。
进一步地，所述的阳离子脂质体材料选自如下至少一种：(2,3-二油氧基丙基)三甲基氯化铵、N-[1-(2,3-二油酰氯)丙基]-N,N,N-氯化三甲胺、阳离子胆固醇、三氟乙酸二甲基-2,3-二油烯氧基丙基-2-(2-精胺甲酰氨基)乙基铵、溴化三甲基十二烷基铵、溴化三甲基十四烷基铵、溴化三甲基十六烷基铵、溴化二甲基双十八烷基铵。
进一步地，所述疫苗剂型为肌肉注射剂、注射剂、滴鼻剂、喷雾剂或吸入剂。
优选地，所述疫苗为滴鼻剂、喷雾剂或吸入剂。
最后，本发明提供了制备上述腺病毒的方法，包括以下步骤：构建含有上述多核苷酸的穿梭质粒载体；然后将构建的穿梭质粒载体与骨架质粒一起转染入宿主细胞，培养宿主细胞；获得复制缺陷重组腺病毒，然后扩大培养、纯化，即可。
进一步地，所述穿梭质粒载体为pDC316-S、pDC315-S、pDC516-S或pDC515-S中至少一种。
进一步地，所述骨架质粒为pBHGlox_E1,3Cre、pBHGlox_E1,3FLP中至少一种。
进一步地，所述宿主细胞为HEK293。
本发明还提供了上述多核苷酸，上述疫苗在制备预防和/或治疗呼吸道病毒感染或致病药物中的用途。优选地，所述呼吸道病毒感染是SARS-CoV-2或其变异株感染引起的。
SEQ ID No.1S蛋白或其突变体自身的信号肽核苷酸序列
ATGTTCGTGTTCCTGGTGCTGCTGCCCCTGGTGAGC。
SEQ ID No.2S蛋白或其突变体自身的信号肽氨基酸序列
MFVFLVLLPLVS。
SEQ ID No.3野生型Ad-S疫苗S抗原的核苷酸序列
ATGTTCGTGTTCCTGGTGCTGCTGCCCCTGGTGAGCAGCCAGTGTGTGAACCTGACAACAAGGACCCAGCTGCCTCCCGCCTACACCAACAGCTTCACAAGGGGCGTGTACTACCCTGACAAGGTGTTCAGGAGCAGCGTGCTGCACTCCACACAGGACCTGTTCCTGCCCTTCTTCTCCAACGTGACATGGTTCCACGCCATCCACGTGTCCGGCACCAACGGCACAAAGAGATTCGACAACCCCGTGCTGCCTTTCAACGACGGCGTGTACTTCGCCTCCACAGAGAAGTC
CAACATCATCAGGGGCTGGATCTTCGGCACAACACTGGACAGCAAGACACAGAGCCTG
CTGATCGTGAACAACGCCACAAACGTGGTCATTAAGGTGTGTGAGTTCCAGTTCTGCAA
CGACCCCTTCCTGGGCGTGTACTATCACAAGAACAACAAGAGCTGGATGGAGAGCGAG
TTCAGGGTGTACTCCAGCGCCAACAACTGTACATTCGAGTACGTGTCCCAGCCTTTCCTG
ATGGACCTGGAGGGCAAGCAGGGCAACTTCAAGAACCTGAGGGAGTTCGTGTTCAAGA
ACATCGACGGCTACTTCAAGATCTACTCCAAGCACACCCCCATCAACCTGGTGAGAGAC
CTGCCTCAGGGCTTCTCCGCCCTGGAGCCTCTGGTGGACCTGCCCATCGGCATCAACAT
CACAAGATTCCAGACACTGCTGGCCCTGCACAGAAGCTACCTGACCCCCGGCGATTCCA
GCAGCGGCTGGACCGCTGGCGCCGCTGCTTACTACGTGGGCTACCTGCAGCCCAGGACA
TTCCTGCTGAAGTACAACGAGAACGGCACAATCACCGATGCCGTGGATTGCGCCCTGGA
TCCTCTGAGCGAGACCAAGTGTACACTGAAGTCCTTCACAGTGGAGAAGGGCATCTACC
AGACCTCCAACTTCAGAGTGCAGCCCACAGAGAGCATCGTGAGATTCCCTAACATCACA
AACCTGTGCCCTTTCGGCGAGGTGTTCAACGCCACAAGGTTCGCCAGCGTGTACGCCTG
GAACAGAAAGAGGATCAGCAACTGCGTGGCCGATTACAGCGTGCTGTACAACTCCGCC
TCCTTCTCCACATTCAAGTGCTACGGCGTGTCCCCCACCAAGCTGAACGATCTGTGTTTC
ACCAACGTGTACGCCGACTCCTTCGTGATCAGAGGCGACGAGGTGAGGCAGATCGCCC
CCGGACAGACCGGCAAGATCGCCGATTACAACTACAAGCTGCCCGATGACTTCACCGGC
TGTGTGATCGCCTGGAACTCCAACAACCTGGATTCCAAGGTGGGCGGCAACTACAACTA
CCTGTACAGACTGTTCAGAAAGTCCAACCTGAAGCCCTTCGAGAGAGATATCTCCACCG
AGATCTACCAGGCCGGCTCCACACCTTGTAACGGCGTGGAGGGCTTCAACTGCTACTTC
CCCCTGCAGAGCTACGGCTTCCAGCCTACCAACGGCGTGGGCTACCAGCCTTACAGAGT
GGTGGTGCTGTCCTTCGAGCTGCTGCACGCCCCCGCCACAGTGTGTGGCCCAAAGAAG
TCCACCAACCTGGTGAAGAACAAGTGTGTGAACTTCAACTTCAACGGCCTGACCGGCA
CAGGCGTGCTGACCGAGAGCAACAAGAAGTTCCTGCCTTTCCAGCAGTTCGGCAGAGA
CATCGCCGATACCACAGACGCCGTGAGAGACCCTCAGACCCTGGAGATCCTGGATATCA
CCCCTTGCAGCTTCGGCGGCGTGTCCGTGATCACCCCTGGCACAAACACCAGCAACCA
GGTGGCCGTGCTGTACCAGGACGTGAACTGTACAGAGGTGCCCGTGGCCATCCACGCC
GACCAGCTGACCCCCACATGGAGAGTGTACTCCACCGGCTCCAACGTGTTCCAGACAA
GGGCCGGCTGCCTGATCGGCGCCGAGCATGTGAACAACAGCTACGAGTGTGACATCCCT
ATCGGCGCCGGCATCTGCGCCAGCTACCAGACCCAGACCAACAGCCCCAGGAGGGCCA
GATCCGTGGCCAGCCAGAGCATCATCGCCTACACCATGTCCCTGGGCGCCGAGAACAGC
GTGGCCTACTCCAACAACAGCATCGCCATCCCCACCAACTTCACAATCTCCGTGACCAC
CGAGATCCTGCCCGTGAGCATGACCAAGACATCCGTGGATTGTACCATGTACATCTGCGG
CGACAGCACAGAGTGCTCCAACCTGCTGCTGCAGTACGGCTCCTTCTGTACCCAGCTGA
ACAGGGCCCTGACAGGCATCGCCGTGGAGCAGGACAAGAACACCCAGGAGGTGTTCG
CCCAGGTGAAGCAGATCTACAAGACACCTCCCATCAAGGACTTCGGCGGCTTCAACTTC
TCCCAGATCCTGCCCGACCCTTCCAAGCCCTCCAAGAGATCCTTCATCGAGGACCTGCT
GTTCAACAAGGTGACCCTGGCCGATGCCGGCTTCATCAAGCAGTACGGCGACTGCCTGG
GCGACATCGCCGCCAGAGACCTGATCTGCGCCCAGAAGTTCAACGGCCTCACCGTGCT
GCCCCCTCTGCTGACCGATGAGATGATCGCCCAGTACACCAGCGCCCTGCTGGCCGGAA
CAATCACCAGCGGCTGGACATTCGGCGCCGGCGCTGCTCTGCAGATCCCTTTCGCCATG
CAGATGGCCTACAGGTTCAACGGCATCGGCGTGACCCAGAACGTGCTGTACGAGAACC
AGAAGCTGATCGCCAACCAGTTCAACAGCGCCATCGGCAAGATCCAGGATTCCCTGAGC
AGCACAGCCAGCGCCCTGGGCAAGCTGCAGGATGTGGTGAACCAGAACGCCCAGGCCC
TGAACACACTGGTGAAGCAGCTGAGCAGCAACTTCGGCGCCATCAGCAGCGTGCTCAA
CGACATCCTGAGCAGACTGGATAAGGTGGAGGCCGAGGTGCAGATCGATAGACTGATCA
CCGGCAGGCTGCAGTCCCTGCAGACATACGTGACCCAGCAGCTGATCAGAGCCGCCGA
GATCAGGGCCAGCGCCAACCTGGCCGCCACCAAGATGTCCGAGTGTGTGCTGGGCCAG
AGCAAGAGGGTGGACTTCTGTGGCAAGGGCTACCACCTGATGTCCTTCCCTCAGTCCGC
CCCCCACGGCGTGGTGTTCCTGCACGTGACATACGTGCCTGCCCAGGAGAAGAACTTCA
CAACAGCCCCTGCCATCTGTCACGACGGCAAGGCCCACTTCCCCAGGGAGGGCGTGTT
CGTGAGCAACGGCACCCACTGGTTCGTGACCCAGAGGAACTTCTACGAGCCTCAGATCA
TCACAACCGATAACACATTCGTGAGCGGCAACTGTGATGTGGTCATTGGCATCGTGAAC
AATACCGTGTACGATCCCCTGCAGCCTGAGCTGGACTCCTTCAAGGAGGAGCTGGATAA
GTACTTCAAGAACCACACCTCCCCCGACGTGGATCTGGGCGACATTAGCGGCATCAACG
CCTCCGTGGTGAACATCCAGAAGGAGATCGACAGGCTGAACGAGGTGGCCAAGAACCT
GAACGAGTCCCTGATCGATCTGCAGGAGCTGGGCAAGTACGAGCAGTACATCAAGTGG
CCCTGGTACATCTGGCTGGGCTTCATCGCCGGCCTGATCGCCATCGTGATGGTGACAATC
ATGCTGTGTTGCATGACATCCTGTTGTTCCTGTCTGAAGGGCTGTTGTAGCTGTGGCTCC
TGTTGCAAGTTCGACGAGGATGACTCCGAGCCCGTGCTGAAGGGCGTGAAGCTGCACT
ACACCTGA
SEQ ID No.4野生型Ad-S疫苗S抗原的氨基酸序列
MFVFLVLLPLVSSQCVNLTTRTQLPPAYTNSFTRGVYYPDKVFRSSVLHSTQDLFLPFFSNVTWFHAIHVSGTNGTKRFDNPVLPFNDGVYFASTEKSNIIRGWIFGTTLDSKTQSLLIVNNATNVVIKVCEFQFCNDPFLGVYYHKNNKSWMESEFRVYSSANNCTFEYVSQPFLMDLEGKQGNFKNLREFVFKNIDGYFKIYSKHTPINLVRDLPQGFSALEPLVDLPIGINITRFQTLLALHRSYLTPGDSSSGWTAGAAAYYVGYLQPRTFLLKYNENGTITDAVDCALDPLSETKCTLKSFTVEKGIYQTSNFRVQPTESIVRFPNITNLCPFGEVFNATRFASVYAWNRKRISNCVADYSVLYNSASFSTFKCYGVSPTKLNDLCFTNVYADSFVIRGDEVRQIAPGQTGKIADYNYKLPDDFTGCVIAWNSNNLDSKVGGNYNYLYRLFRKSNLKPFERDISTEIYQAGSTPCNGVEGFNCYFPLQSYGFQPTNGVGYQPYRVVVLSFELLHAPATVCGPKKSTNLVKNKCVNFNFNGLTGTGVLTESNKKFLPFQQFGRDIADTTDAVRDPQTLEILDITPCSFGGVSVITPGTNTSNQVAVLYQDVNCTEVPVAIHADQLTPTWRVYSTGSNVFQTRAGCLIGAEHVNNSYECDIPIGAGICASYQTQTNSPRRARSVASQSIIAYTMSLGAENSVAYSNNSIAIPTNFTISVTTEILPVSMTKTSVDCTMYICGDSTECSNLLLQYGSFCTQLNRALTGIAVEQDKNTQEVFAQVKQIYKTPPIKDFGGFNFSQILPDPSKPSKRSFIEDLLFNKVTLADAGFIKQYGDCLGDIAARDLICAQKFNGLTVLPPLLTDEMIAQYTSALLAGTITSGWTFGAGAALQIPFAMQMAYRFNGIGVTQNVLYENQKLIANQFNSAIGKIQDSLSSTASALGKLQDVVNQNAQALNTLVKQLSSNFGAISSVLNDILSRLDKVEAEVQIDRLITGRLQSLQTYVTQQLIRAAEIRASANLAATKMSECVLGQSKRVDFCGKGYHLMSFPQSAPHGVVFLHVTYVPAQEKNFTTAPAICHDGKAHFPREGVFVSNGTHWFVTQRNFYEPQIITTDNTFVSGNCDVVIGIVNNTVYDPLQPELDSFKEELDKYFKNHTSPDVDLGDISGINASVVNIQKEIDRLNEVAKNLNESLIDLQELGKYEQYIKWPWYIWLGFIAGLIAIVMVTIMLCCMTSCCSCLKGCCSCGSCCKFDEDDSEPVLKGVKLHYT*
SEQ ID No.6、SEQ ID No.8、SEQ ID No.10、SEQ ID No.12、SEQ ID No.14、SEQ IDNo.16或SEQ ID No.18是将SEQ ID No.4的S蛋白的第986位赖氨酸(K)替换为脯氨酸(P)，将987位缬氨酸(V)替换为脯氨酸(P)。
SEQ ID No.5、SEQ ID No.7、SEQ ID No.9、SEQ ID No.11、SEQ ID No.13、SEQ IDNo.15或SEQ ID No.17是在野生型S蛋白基因的核苷酸序列基础上结合各变异株S蛋白突变位点的氨基酸得到各变异株S蛋白基因的核苷酸序列，并进行密码子优化而得。
SEQ ID No.5Ad-S-Beta疫苗S抗原的核苷酸序列
ATGTTCGTGTTCCTGGTGCTGCTGCCCCTGGTGAGCAGCCAATGCGTGAACCTGACCACAAGAACACAGCTGCCCCCCGCCTACACCAACAGCTTCACAAGAGGCGTGTATTATCCCGACAAGGTGTTCAGAAGCAGCGTGCTGCACAGCACCCAAGACCTGTTTCTGCCTTTT
TTCAGCAACGTGACCTGGTTCCACGCCATCCACGTGAGCGGCACCAACGGCACCAAGA
GATTCGCCAACCCCGTGCTGCCCTTCAACGACGGCGTGTACTTCGCTAGCACCGAGAAG
AGCAACATCATCAGAGGCTGGATCTTCGGCACCACCCTGGATAGCAAGACACAGAGCCT
GCTGATCGTGAACAACGCCACCAACGTGGTGATCAAGGTGTGCGAGTTTCAGTTCTGCA
ACGACCCCTTCCTGGGCGTGTATTACCACAAGAACAACAAGAGCTGGATGGAGAGCGA
GTTCAGAGTCTACAGCAGCGCCAACAACTGCACCTTCGAGTACGTGAGCCAACCCTTCC
TGATGGACCTGGAGGGCAAGCAAGGCAATTTCAAGAACCTGAGAGAGTTCGTGTTCAA
GAACATCGACGGCTACTTCAAGATCTACAGCAAGCACACCCCCATCAACCTGGTGAGAG
GCCTGCCCCAAGGCTTCAGCGCCCTGGAGCCCCTGGTGGACCTGCCCATCGGCATCAAC
ATCACAAGATTTCAGACACTGCACCGGTCCTATCTCACCCCTGGCGACAGCAGCAGCGG
GTGGACCGCTGGCGCCGCCGCTTACTACGTGGGCTACCTGCAGCCTAGAACCTTCCTGC
TGAAGTACAACGAGAACGGCACCATTACCGACGCCGTGGACTGCGCCCTGGACCCCCT
GAGCGAGACCAAGTGCACCCTGAAGAGCTTCACCGTGGAGAAGGGCATCTATCAGACA
AGCAACTTCAGAGTGCAGCCCACCGAGAGCATCGTGAGATTCCCCAACATCACCAACCT
GTGCCCCTTCGGCGAGGTGTTCAACGCCACAAGATTCGCTAGCGTGTACGCCTGGAACC
GGAAAAGAATCAGCAACTGCGTGGCCGACTACAGCGTGCTGTACAACAGCGCTAGCTT
CAGCACCTTCAAGTGCTACGGCGTGAGCCCCACCAAGCTGAACGACCTGTGCTTCACC
AACGTGTACGCCGACAGCTTCGTGATCAGAGGCGACGAGGTGAGACAGATCGCCCCCG
GGCAGACCGGCAACATCGCCGACTACAACTACAAGCTGCCCGACGACTTCACCGGCTG
CGTGATCGCCTGGAACAGCAATAACCTGGACAGCAAGGTGGGCGGCAACTACAACTAC
CTGTACAGACTGTTCAGAAAGAGCAACCTGAAGCCCTTCGAGAGAGACATCAGCACCG
AGATCTACCAAGCCGGCAGCACCCCCTGCAACGGCGTGAAGGGCTTCAACTGCTACTTC
CCCCTGCAGAGCTACGGCTTTCAGCCCACCTACGGCGTGGGCTATCAGCCCTACAGAGT
GGTCGTGCTGAGCTTCGAGCTGCTGCACGCCCCTGCCACCGTGTGTGGCCCCAAGAAG
AGCACCAACCTGGTGAAGAACAAGTGCGTGAACTTCAACTTCAACGGGCTGACCGGGA
CCGGCGTGCTGACCGAGAGCAACAAGAAGTTTCTGCCCTTTCAACAGTTCGGCAGAGA
CATCGCCGACACCACCGACGCCGTCCGGGACCCTCAGACCCTGGAGATCCTGGACATCA
CCCCCTGCTCCTTCGGCGGCGTGAGCGTGATCACCCCCGGCACCAACACAAGCAACCA
AGTGGCCGTGCTGTACCAAGGCGTGAACTGCACCGAGGTGCCCGTGGCCATCCACGCC
GATCAGCTGACCCCCACCTGGAGAGTGTACAGCACCGGCAGCAACGTGTTTCAGACAA
GAGCCGGCTGCCTGATCGGCGCCGAGCACGTGAACAACAGCTACGAGTGCGACATCCC
CATCGGCGCCGGCATCTGCGCTAGCTATCAGACACAGACCAACAGCCCTAGAAGAGCTA
GAAGCGTGGCTAGCCAAAGCATCATCGCCTACACCATGAGCCTGGGCGTGGAGAACAG
CGTGGCCTACAGCAACAACAGCATCGCCATCCCCACCAACTTCACCATCAGCGTGACCA
CCGAAATCCTGCCCGTGAGCATGACCAAGACAAGCGTGGACTGCACCATGTACATCTGC
GGCGACAGCACCGAGTGCAGCAACCTGCTCCTGCAGTACGGCAGCTTCTGCACACAGC
TGAACAGAGCCCTGACCGGCATCGCCGTGGAGCAAGACAAGAACACCCAAGAGGTGTT
CGCCCAAGTGAAGCAGATCTACAAGACCCCCCCCATCAAGGACTTCGGCGGCTTCAACT
TCAGCCAAATTCTGCCCGACCCTAGCAAGCCTAGCAAGAGAAGCTTCATCGAGGACCTG
CTGTTCAACAAGGTGACCCTGGCCGACGCCGGCTTCATCAAGCAGTACGGCGACTGCCT
GGGCGATATCGCTGCTAGAGACCTGATCTGCGCTCAGAAGTTCAACGGCCTGACCGTGC
TCCCCCCCCTGCTGACCGACGAGATGATCGCTCAGTACACAAGCGCTCTGCTCGCCGGG
ACCATCACATCCGGGTGGACATTCGGGGCCGGCGCTGCCCTGCAGATCCCCTTCGCCAT
GCAGATGGCCTACAGATTCAACGGCATCGGCGTGACACAGAACGTGCTGTACGAGAATC
AGAAGCTGATCGCCAATCAGTTCAACAGCGCCATCGGCAAGATCCAAGACAGCCTGAG
CAGCACCGCTAGCGCCCTGGGCAAGCTGCAAGACGTGGTGAATCAGAACGCCCAAGCC
CTGAACACCCTGGTGAAGCAGCTGAGCAGCAACTTCGGCGCCATCAGCTCCGTGCTGA
ACGACATCCTGAGCAGACTGGACCCCCCCGAGGCCGAGGTGCAGATTGACAGACTGAT
CACCGGCAGACTGCAGAGCCTGCAGACCTACGTGACACAGCAGCTGATCAGAGCCGCC
GAGATCAGAGCTAGCGCCAACCTGGCCGCCACCAAGATGAGCGAGTGCGTGCTGGGGC
AGAGCAAGAGAGTGGACTTCTGCGGCAAGGGCTACCACCTGATGAGCTTCCCTCAGAG
CGCCCCCCACGGCGTGGTGTTCCTGCACGTGACCTACGTGCCCGCCCAAGAGAAGAAC
TTCACCACCGCCCCCGCCATCTGCCACGACGGCAAGGCCCACTTCCCTAGAGAGGGCGT
GTTCGTGAGCAACGGCACCCACTGGTTCGTGACACAGAGAAACTTCTACGAGCCTCAG
ATCATCACCACCGACAACACCTTCGTGAGCGGCAACTGCGACGTGGTGATCGGCATTGT
GAACAATACCGTGTACGACCCCCTGCAGCCCGAGCTGGACAGCTTCAAGGAGGAGCTG
GACAAGTACTTCAAGAACCACACAAGCCCCGACGTGGACCTGGGCGACATTAGCGGCA
TCAACGCTAGCGTGGTGAACATTCAGAAGGAGATCGACAGACTGAACGAGGTGGCCAA
GAACCTGAACGAGAGCCTGATCGACCTGCAAGAGCTGGGCAAGTACGAGCAGTACATC
AAGTGGCCCTGGTACATCTGGCTGGGCTTCATCGCCGGCCTGATCGCCATCGTGATGGTG
ACCATCATGCTGTGCTGCATGACAAGCTGTTGCAGCTGCCTGAAGGGCTGCTGCAGCTG
CGGGAGCTGCTGCAAGTTCGACGAGGACGACAGCGAGCCCGTGCTGAAGGGCGTGAA
GCTGCACTACACCTGA
SEQ ID No.6Ad-S-Beta疫苗S抗原的氨基酸序列
MFVFLVLLPLVSSQCVNLTTRTQLPPAYTNSFTRGVYYPDKVFRSSVLHSTQDLFLPFFSNVTWFHAIHVSGTNGTKRFANPVLPFNDGVYFASTEKSNIIRGWIFGTTLDSKTQSLLIVNNATNVVIKVCEFQFCNDPFLGVYYHKNNKSWMESEFRVYSSANNCTFEYVSQPFLMDLEGKQGNFKNLREFVFKNIDGYFKIYSKHTPINLVRGLPQGFSALEPLVDLPIGINITRFQTLHRSYLTPGDSSSGWTAGAAAYYVGYLQPRTFLLKYNENGTITDAVDCALDPLSETKCTLKSFTVEKGIYQTSNFRVQPTESIVRFPNITNLCPFGEVFNATRFASVYAWNRKRISNCVADYSVLYNSASFSTFKCYGVSPTKLNDLCFTNVYADSFVIRGDEVRQIAPGQTGNIADYNYKLPDDFTGCVIAWNSNNLDSKVGGNYNYLYRLFRKSNLKPFERDISTEIYQAGSTPCNGVKGFNCYFPLQSYGFQPTYGVGYQPYRVVVLSFELLHAPATVCGPKKSTNLVKNKCVNFNFNGLTGTGVLTESNKKFLPFQQFGRDIADTTDAVRDPQTLEILDITPCSFGGVSVITPGTNTSNQVAVLYQGVNCTEVPVAIHADQLTPTWRVYSTGSNVFQTRAGCLIGAEHVNNSYECDIPIGAGICASYQTQTNSPRRARSVASQSIIAYTMSLGVENSVAYSNNSIAIPTNFTISVTTEILPVSMTKTSVDCTMYICGDSTECSNLLLQYGSFCTQLNRALTGIAVEQDKNTQEVFAQVKQIYKTPPIKDFGGFNFSQILPDPSKPSKRSFIEDLLFNKVTLADAGFIKQYGDCLGDIAARDLICAQKFNGLTVLPPLLTDEMIAQYTSALLAGTITSGWTFGAGAALQIPFAMQMAYRFNGIGVTQNVLYENQKLIANQFNSAIGKIQDSLSSTASALGKLQDVVNQNAQALNTLVKQLSSNFGAISSVLNDILSRLDPPEAEVQIDRLITGRLQSLQTYVTQQLIRAAEIRASANLAATKMSECVLGQSKRVDFCGKGYHLMSFPQSAPHGVVFLHVTYVPAQEKNFTTAPAICHDGKAHFPREGVFVSNGTHWFVTQRNFYEPQIITTDNTFVSGNCDVVIGIVNNTVYDPLQPELDSFKEELDKYFKNHTSPDVDLGDISGINASVVNIQKEIDRLNEVAKNLNESLIDLQELGKYEQYIKWPWYIWLGFIAGLIAIVMVTIMLCCMTSCCSCLKGCCSCGSCCKFDEDDSEPVLKGVKLHYT*
SEQ ID No.7Ad-S-Delta疫苗S抗原的核苷酸序列
ATGTTCGTGTTCCTGGTGCTGCTGCCCCTGGTGAGCAGCCAATGCGTGAACCTGAGAACAAGAACACAGCTGCCCCCCGCCTACACCAACAGCTTCACAAGAGGGGTCTACTACCCCGACAAGGTGTTCAGAAGCAGCGTGCTGCACAGCACCCAAGACCTGTTCCTGCCCTTCTTCTCCAACGTGACCTGGTTCCACGCCATCCACGTGAGCGGCACCAACGGCACCAAGAGATTCGACAACCCCGTGCTGCCCTTCAACGACGGCGTGTACTTCGCTAGCACCGAGAAGAGCAACATCATCAGAGGCTGGATCTTCGGCACCACCCTGGACAGCAAGACACAGAGC
CTGCTGATTGTGAACAACGCTACCAACGTGGTGATCAAGGTGTGCGAGTTTCAGTTCTG
CAACGACCCCTTCCTGGGGGTGTACTACCACAAAAACAACAAGAGCTGGATGGAGAGC
GGCGTGTACAGCAGCGCCAACAACTGCACCTTCGAGTACGTGAGCCAACCCTTCCTGAT
GGACCTGGAGGGCAAGCAAGGCAACTTCAAGAACCTCAGAGAGTTCGTGTTCAAGAA
CATCGACGGCTACTTCAAGATCTACAGCAAGCACACCCCCATCAACCTGGTGAGAGACC
TGCCCCAAGGCTTCAGCGCCCTGGAGCCCCTGGTGGACCTGCCCATCGGCATCAACATC
ACAAGATTTCAGACCCTGCTGGCCCTGCACAGAAGCTACCTGACACCCGGCGACAGCA
GCAGCGGGTGGACAGCCGGCGCTGCCGCTTACTACGTGGGCTACCTGCAGCCTAGAAC
CTTCCTGCTGAAGTACAACGAGAACGGCACCATTACCGACGCCGTCGACTGCGCCCTGG
ACCCCCTGAGCGAGACCAAGTGCACCCTGAAGAGCTTCACCGTGGAGAAGGGCATCTA
TCAGACAAGCAACTTCAGAGTGCAGCCCACCGAGAGCATCGTGAGATTCCCCAACATC
ACCAACCTGTGCCCCTTCGGCGAGGTGTTCAACGCCACAAGATTCGCTAGCGTGTACGC
CTGGAACAGAAAGAGAATCAGCAACTGCGTGGCCGACTACAGCGTGCTGTACAACAGC
GCTAGCTTCAGCACCTTCAAATGCTACGGGGTCAGCCCCACCAAGCTGAACGACCTGTG
CTTCACCAACGTGTACGCCGACAGCTTCGTGATCAGAGGCGACGAGGTGAGACAGATC
GCCCCCGGGCAGACCGGCAAGATCGCCGACTACAACTACAAGCTGCCCGACGACTTCA
CCGGCTGCGTGATCGCCTGGAACTCCAACAACCTGGATAGCAAGGTGGGCGGCAACTA
CAACTATAGATACAGACTGTTCAGAAAGAGCAACCTGAAGCCCTTCGAGAGAGACATCA
GCACCGAGATCTACCAAGCCGGCAGCAAGCCCTGCAACGGCGTGGAGGGCTTCAACTG
CTACTTCCCCCTGCAGAGCTACGGCTTTCAGCCCACCAACGGCGTGGGCTATCAGCCCT
ACAGAGTGGTCGTGCTGAGCTTCGAGCTGCTGCACGCCCCTGCCACCGTGTGCGGCCC
CAAGAAGAGCACCAACCTGGTGAAGAACAAGTGCGTGAACTTCAACTTCAACGGCCTG
ACCGGCACCGGCGTGCTGACCGAGAGCAACAAGAAGTTCCTCCCTTTTCAGCAGTTCG
GCAGAGACATCGCCGACACCACCGATGCCGTGAGAGACCCTCAGACCCTGGAGATCCT
GGACATCACCCCCTGCAGCTTTGGCGGCGTGAGCGTGATCACCCCCGGCACCAACACA
AGCAACCAAGTGGCCGTGCTGTACCAAGGCGTGAACTGCACCGAGGTGCCCGTGGCCA
TCCACGCCGATCAGCTGACCCCCACCTGGAGAGTGTACAGCACCGGCAGCAACGTGTTT
CAGACAAGAGCCGGCTGCCTGATCGGCGCCGAGCACGTGAACAACAGCTACGAGTGCG
ACATCCCCATCGGCGCCGGCATCTGCGCTAGCTATCAGACACAGACCAACAGCAGACGG
AGAGCTAGAAGCGTGGCTAGCCAAAGCATCATCGCCTACACCATGAGCCTGGGCGCCGA
GAACAGCGTGGCCTACAGCAACAACAGCATCGCCATCCCCACCAACTTCACCATCAGCG
TGACCACCGAGATCCTGCCCGTCTCCATGACCAAGACAAGCGTGGACTGCACCATGTAC
ATCTGCGGCGACAGCACCGAGTGCAGCAACCTGCTCCTGCAGTACGGCAGCTTCTGCAC
ACAGCTGAACAGAGCCCTGACCGGCATCGCCGTGGAGCAAGACAAGAACACCCAAGA
GGTGTTCGCCCAAGTGAAGCAGATCTACAAGACCCCCCCCATCAAGGACTTCGGCGGCT
TCAACTTCAGCCAAATCCTGCCCGACCCTAGCAAGCCTAGCAAGAGAAGCTTCATCGAG
GACCTGCTGTTCAACAAGGTGACCCTGGCCGACGCCGGCTTCATCAAGCAGTACGGCG
ACTGCCTGGGCGACATCGCCGCTAGAGACCTGATCTGCGCTCAGAAGTTCAATGGCCTG
ACCGTCCTGCCCCCCCTGCTGACCGACGAGATGATCGCTCAGTACACAAGCGCCCTGCT
GGCCGGCACAATTACAAGCGGCTGGACATTCGGCGCTGGCGCCGCCCTGCAAATCCCCT
TCGCCATGCAGATGGCCTACAGATTCAACGGCATCGGCGTGACACAGAACGTGCTGTAC
GAGAATCAGAAGCTGATCGCCAATCAGTTCAACAGCGCCATCGGCAAGATCCAAGACA
GCCTGAGCAGCACCGCTAGCGCCCTGGGCAAGCTGCAGAACGTGGTGAATCAGAACGC
CCAAGCCCTGAACACCCTGGTGAAGCAGCTGAGCAGCAACTTCGGCGCCATCAGCTCC
GTGCTGAACGATATCCTGAGCAGACTGGACCCCCCCGAGGCCGAGGTGCAGATCGACC
GGCTGATCACCGGCAGACTGCAGAGCCTGCAGACCTACGTGACACAGCAGCTGATCAG
AGCCGCCGAGATCAGAGCTAGCGCCAACCTGGCCGCCACCAAGATGAGCGAGTGCGTG
CTGGGGCAGAGCAAGAGAGTGGACTTCTGCGGCAAGGGCTACCACCTGATGAGCTTCC
CTCAGAGCGCCCCCCACGGCGTGGTGTTCCTGCACGTGACCTACGTGCCCGCCCAAGA
GAAGAACTTCACCACCGCCCCCGCCATCTGCCACGACGGCAAGGCCCACTTCCCTAGA
GAGGGCGTGTTCGTGAGCAACGGCACCCACTGGTTCGTGACACAGAGAAACTTCTACG
AGCCTCAGATCATCACCACCGACAACACCTTCGTGAGCGGCAACTGCGACGTGGTGATC
GGCATTGTGAACAATACAGTCTACGACCCTCTGCAGCCCGAGCTGGACAGCTTCAAGGA
GGAGCTGGACAAGTACTTCAAGAATCACACAAGCCCCGACGTGGACCTGGGGGACATC
AGCGGCATCAACGCTAGCGTGGTGAACATTCAGAAGGAGATCGACAGACTGAACGAGG
TGGCCAAGAACCTGAACGAGAGCCTGATCGACCTGCAAGAGCTGGGCAAGTACGAGCA
GTACATCAAGTGGCCCTGGTACATCTGGCTGGGCTTCATCGCCGGCCTGATCGCCATCGT
GATGGTGACCATCATGCTGTGCTGCATGACAAGCTGCTGCAGCTGCCTGAAGGGCTGCT
GCTCCTGCGGCAGCTGCTGCAAGTTCGACGAGGACGACAGCGAGCCCGTGCTGAAGGG
CGTGAAGCTGCACTACACCTGA
SEQ ID No.8Ad-S-Delta疫苗S抗原的氨基酸序列
MFVFLVLLPLVSSQCVNLRTRTQLPPAYTNSFTRGVYYPDKVFRSSVLHSTQDLFLPFFSNVTWFHAIHVSGTNGTKRFDNPVLPFNDGVYFASTEKSNIIRGWIFGTTLDSKTQSLLIVNNATNVVIKVCEFQFCNDPFLGVYYHKNNKSWMESGVYSSANNCTFEYVSQPFLMDLEGKQGNFKNLREFVFKNIDGYFKIYSKHTPINLVRDLPQGFSALEPLVDLPIGINITRFQTLLALHRSYLTPGDSSSGWTAGAAAYYVGYLQPRTFLLKYNENGTITDAVDCALDPLSETKCTLKSFTVEKGIYQTSNFRVQPTESIVRFPNITNLCPFGEVFNATRFASVYAWNRKRISNCVADYSVLYNSASFSTFKCYGVSPTKLNDLCFTNVYADSFVIRGDEVRQIAPGQTGKIADYNYKLPDDFTGCVIAWNSNNLDSKVGGNYNYRYRLFRKSNLKPFERDISTEIYQAGSKPCNGVEGFNCYFPLQSYGFQPTNGVGYQPYRVVVLSFELLHAPATVCGPKKSTNLVKNKCVNFNFNGLTGTGVLTESNKKFLPFQQFGRDIADTTDAVRDPQTLEILDITPCSFGGVSVITPGTNTSNQVAVLYQGVNCTEVPVAIHADQLTPTWRVYSTGSNVFQTRAGCLIGAEHVNNSYECDIPIGAGICASYQTQTNSRRRARSVASQSIIAYTMSLGAENSVAYSNNSIAIPTNFTISVTTEILPVSMTKTSVDCTMYICGDSTECSNLLLQYGSFCTQLNRALTGIAVEQDKNTQEVFAQVKQIYKTPPIKDFGGFNFSQILPDPSKPSKRSFIEDLLFNKVTLADAGFIKQYGDCLGDIAARDLICAQKFNGLTVLPPLLTDEMIAQYTSALLAGTITSGWTFGAGAALQIPFAMQMAYRFNGIGVTQNVLYENQKLIANQFNSAIGKIQDSLSSTASALGKLQNVVNQNAQALNTLVKQLSSNFGAISSVLNDILSRLDPPEAEVQIDRLITGRLQSLQTYVTQQLIRAAEIRASANLAATKMSECVLGQSKRVDFCGKGYHLMSFPQSAPHGVVFLHVTYVPAQEKNFTTAPAICHDGKAHFPREGVFVSNGTHWFVTQRNFYEPQIITTDNTFVSGNCDVVIGIVNNTVYDPLQPELDSFKEELDKYFKNHTSPDVDLGDISGINASVVNIQKEIDRLNEVAKNLNESLIDLQELGKYEQYIKWPWYIWLGFIAGLIAIVMVTIMLCCMTSCCSCLKGCCSCGSCCKFDEDDSEPVLKGVKLHYT*
SEQ ID No.9Ad-S-Beta-Delta疫苗S抗原的核苷酸序列
ATGTTCGTGTTCCTGGTGCTGCTGCCCCTGGTGAGCAGCCAATGCGTGAACCTGACCACAAGAACACAGCTGCCCCCCGCCTACACCAACAGCTTCACAAGAGGCGTGTATTATCCCGACAAGGTGTTCAGAAGCAGCGTGCTGCACAGCACCCAAGACCTGTTTCTGCCCTTCTTCAGCAACGTGACCTGGTTCCACGCCATCCACGTGAGCGGCACCAACGGCACCAAGAGATTCGCCAACCCCGTGCTGCCCTTCAACGACGGCGTGTACTTCGCTAGCACCGAGAAGAGCAACATCATCAGAGGCTGGATCTTCGGCACCACCCTGGATAGCAAGACACAGAGCCTGCTGATCGTGAACAACGCCACCAACGTGGTGATCAAGGTGTGCGAGTTTCAGTTCTGCAACGACCCCTTCCTGGGCGTGTATTACCACAAGAACAACAAGAGCTGGATGGAGAGCGAGTTCAGAGTCTACAGCAGCGCCAACAACTGCACCTTCGAGTACGTGAGCCAACCCT
TCCTGATGGACCTGGAGGGCAAGCAAGGCAATTTCAAGAACCTGAGAGAGTTCGTGTT
CAAGAACATCGACGGCTACTTCAAGATCTACAGCAAGCACACCCCCATCAACCTGGTGA
GAGGCCTGCCCCAAGGCTTCAGCGCCCTGGAGCCCCTGGTGGACCTGCCCATCGGCATC
AACATCACAAGATTTCAGACCCTGCACCGGAGCTATCTGACCCCTGGCGACTCCTCCTC
CGGGTGGACCGCTGGCGCCGCCGCTTACTACGTGGGCTACCTGCAGCCTAGAACCTTCC
TGCTGAAGTACAACGAGAACGGCACCATTACAGACGCCGTGGATTGCGCCCTGGACCC
CCTGAGCGAGACCAAGTGCACCCTGAAGAGCTTCACCGTGGAGAAGGGCATCTATCAG
ACAAGCAACTTCAGAGTGCAGCCCACCGAGAGCATCGTGAGATTCCCCAACATCACCA
ACCTGTGCCCCTTCGGCGAGGTGTTCAACGCCACAAGATTCGCTAGCGTGTACGCCTGG
AACAGAAAAAGAATCAGCAACTGCGTGGCCGACTACAGCGTGCTGTACAACAGCGCTA
GCTTCAGCACCTTCAAGTGCTACGGCGTGAGCCCCACCAAGCTGAACGACCTGTGCTTC
ACCAACGTGTACGCCGACAGCTTCGTGATCAGAGGCGACGAGGTGAGACAGATCGCCC
CCGGGCAGACCGGCAACATCGCCGACTACAACTATAAGCTCCCCGACGACTTCACCGGC
TGCGTGATCGCTTGGAACTCCAACAACCTGGACAGCAAGGTGGGCGGCAACTACAACT
ACAGATATAGACTGTTCAGAAAGAGCAACCTGAAGCCCTTCGAGAGAGACATCAGCAC
CGAGATCTACCAAGCCGGCAGCAAGCCCTGCAACGGCGTGAAGGGCTTCAACTGCTAC
TTCCCCCTGCAGAGCTACGGCTTTCAGCCCACCTACGGCGTGGGCTATCAGCCCTACAG
AGTGGTCGTGCTGAGCTTCGAGCTGCTGCACGCCCCCGCCACCGTCTGCGGCCCCAAG
AAGAGCACCAACCTGGTGAAGAACAAGTGCGTGAACTTCAACTTCAACGGGCTGACCG
GCACCGGCGTGCTGACCGAGAGCAACAAGAAGTTCCTGCCCTTTCAGCAGTTCGGCAG
AGACATCGCCGACACCACCGACGCCGTGAGAGACCCTCAGACCCTGGAGATCCTGGAC
ATCACACCCTGCAGCTTCGGCGGCGTGAGCGTGATCACCCCCGGCACCAACACAAGCA
ACCAAGTGGCCGTGCTGTACCAAGGCGTGAACTGCACCGAGGTGCCCGTGGCCATCCA
CGCCGATCAGCTGACCCCCACCTGGAGAGTCTATAGCACCGGCAGCAACGTGTTTCAGA
CAAGAGCCGGCTGCCTGATCGGCGCCGAGCACGTGAACAACAGCTACGAGTGCGACAT
CCCCATCGGCGCCGGCATCTGCGCTAGCTATCAGACACAGACCAACAGCCCTAGAAGAG
CTAGAAGCGTGGCTAGCCAAAGCATCATCGCCTACACCATGAGCCTGGGCGTGGAGAAC
AGCGTGGCCTACAGCAACAACAGCATCGCCATCCCCACCAACTTCACCATCAGCGTGAC
CACCGAGATCCTCCCTGTGAGCATGACCAAGACAAGCGTGGACTGCACCATGTACATCT
GCGGCGACAGCACCGAGTGCAGCAACCTGCTCCTGCAGTACGGCAGCTTCTGCACACA
GCTGAACAGAGCCCTGACCGGCATCGCCGTGGAGCAAGACAAGAACACCCAAGAGGT
GTTCGCCCAAGTGAAGCAGATCTACAAGACCCCCCCCATCAAGGACTTCGGCGGCTTCA
ACTTCAGCCAAATCCTGCCCGACCCTAGCAAGCCTAGCAAGAGAAGCTTCATCGAGGAC
CTGCTGTTCAACAAGGTGACCCTGGCCGACGCCGGCTTCATCAAGCAGTACGGCGACTG
CCTGGGCGACATCGCCGCTAGAGACCTGATCTGCGCTCAGAAGTTCAACGGCCTGACCG
TGCTGCCCCCCCTGCTGACCGACGAGATGATCGCTCAATACACAAGCGCCCTCCTGGCC
GGGACAATCACATCCGGCTGGACCTTCGGCGCCGGGGCTGCCCTGCAGATCCCCTTCGC
CATGCAGATGGCCTACAGATTCAACGGCATCGGCGTGACACAGAACGTGCTGTACGAGA
ATCAGAAGCTGATCGCCAATCAGTTCAACAGCGCCATCGGCAAGATCCAAGACAGCCTG
AGCAGCACCGCTAGCGCCCTGGGCAAGCTGCAAGACGTGGTGAATCAGAACGCCCAAG
CCCTGAACACCCTGGTGAAGCAGCTGAGCAGCAACTTCGGCGCCATCAGCAGCGTGCT
GAACGACATCCTGAGCAGACTGGACCCCCCCGAGGCCGAGGTGCAGATCGATAGACTG
ATCACCGGCAGACTGCAGAGCCTGCAGACCTACGTGACACAGCAGCTGATCAGAGCCG
CCGAGATCAGAGCTAGCGCCAACCTGGCCGCCACCAAGATGAGCGAGTGCGTGCTGGG
GCAGAGCAAGAGAGTGGACTTCTGCGGCAAGGGCTACCACCTGATGAGCTTCCCTCAG
AGCGCCCCCCACGGCGTGGTGTTCCTGCACGTGACCTACGTGCCCGCCCAAGAGAAGA
ACTTCACCACCGCTCCCGCCATCTGCCACGACGGCAAGGCCCACTTCCCTAGAGAGGGC
GTGTTCGTGAGCAACGGCACCCACTGGTTCGTGACACAGAGAAACTTCTACGAGCCTC
AGATCATCACCACCGACAACACCTTCGTGAGCGGCAACTGCGACGTGGTGATCGGCATC
GTGAACAATACCGTCTACGACCCCCTGCAACCCGAGCTGGACAGCTTCAAGGAGGAGC
TGGACAAGTACTTCAAGAACCACACAAGCCCCGACGTGGACCTGGGCGATATCAGCGG
CATCAACGCTAGCGTGGTGAACATTCAGAAGGAAATCGATCGGCTGAATGAGGTGGCCA
AGAACCTGAACGAGAGCCTGATCGACCTGCAAGAGCTGGGCAAGTACGAGCAGTACAT
CAAGTGGCCCTGGTACATCTGGCTGGGCTTCATCGCCGGCCTGATCGCCATCGTGATGGT
GACCATCATGCTGTGCTGCATGACAAGCTGCTGCAGCTGCCTGAAGGGCTGCTGTAGCT
GTGGCAGCTGCTGCAAGTTCGACGAGGACGACAGCGAGCCCGTGCTGAAGGGCGTGA
AGCTGCACTACACCTGA
SEQ ID No.10Ad-S-Beta-Delta疫苗S抗原的氨基酸序列
MFVFLVLLPLVSSQCVNLTTRTQLPPAYTNSFTRGVYYPDKVFRSSVLHSTQDLFLPFFSNVTWFHAIHVSGTNGTKRFANPVLPFNDGVYFASTEKSNIIRGWIFGTTLDSKTQSLLIVNNATNVVIKVCEFQFCNDPFLGVYYHKNNKSWMESEFRVYSSANNCTFEYVSQPFLMDLEGKQGNFKNLREFVFKNIDGYFKIYSKHTPINLVRGLPQGFSALEPLVDLPIGINITRFQTLHRSYLTPGDSSSGWTAGAAAYYVGYLQPRTFLLKYNENGTITDAVDCALDPLSETKCTLKSFTVEKGIYQTSNFRVQPTESIVRFPNITNLCPFGEVFNATRFASVYAWNRKRISNCVADYSVLYNSASFSTFKCYGVSPTKLNDLCFTNVYADSFVIRGDEVRQIAPGQTGNIADYNYKLPDDFTGCVIAWNSNNLDSKVGGNYNYRYRLFRKSNLKPFERDISTEIYQAGSKPCNGVKGFNCYFPLQSYGFQPTYGVGYQPYRVVVLSFELLHAPATVCGPKKSTNLVKNKCVNFNFNGLTGTGVLTESNKKFLPFQQFGRDIADTTDAVRDPQTLEILDITPCSFGGVSVITPGTNTSNQVAVLYQGVNCTEVPVAIHADQLTPTWRVYSTGSNVFQTRAGCLIGAEHVNNSYECDIPIGAGICASYQTQTNSPRRARSVASQSIIAYTMSLGVENSVAYSNNSIAIPTNFTISVTTEILPVSMTKTSVDCTMYICGDSTECSNLLLQYGSFCTQLNRALTGIAVEQDKNTQEVFAQVKQIYKTPPIKDFGGFNFSQILPDPSKPSKRSFIEDLLFNKVTLADAGFIKQYGDCLGDIAARDLICAQKFNGLTVLPPLLTDEMIAQYTSALLAGTITSGWTFGAGAALQIPFAMQMAYRFNGIGVTQNVLYENQKLIANQFNSAIGKIQDSLSSTASALGKLQDVVNQNAQALNTLVKQLSSNFGAISSVLNDILSRLDPPEAEVQIDRLITGRLQSLQTYVTQQLIRAAEIRASANLAATKMSECVLGQSKRVDFCGKGYHLMSFPQSAPHGVVFLHVTYVPAQEKNFTTAPAICHDGKAHFPREGVFVSNGTHWFVTQRNFYEPQIITTDNTFVSGNCDVVIGIVNNTVYDPLQPELDSFKEELDKYFKNHTSPDVDLGDISGINASVVNIQKEIDRLNEVAKNLNESLIDLQELGKYEQYIKWPWYIWLGFIAGLIAIVMVTIMLCCMTSCCSCLKGCCSCGSCCKFDEDDSEPVLKGVKLHYT*
SEQ ID No.11Ad-S-Omicron-BA.1疫苗S抗原的核苷酸序列
ATGTTCGTGTTCCTGGTGCTGCTGCCCCTGGTGAGCAGCCAATGCGTGAACCTGACCACAAGAACACAGCTGCCCCCCGCCTACACCAACAGCTTCACAAGAGGCGTGTACTACCCCGACAAGGTGTTCAGAAGCAGCGTCCTCCACAGCACCCAAGACCTGTTCCTGCCCTTCTTCAGCAACGTGACCTGGTTCCACGTGATCAGCGGCACCAACGGCACCAAGAGATTCGACAACCCCGTGCTGCCCTTCAACGACGGCGTGTACTTCGCTAGCATCGAGAAGAGCAACATCATCAGAGGCTGGATCTTCGGCACCACCCTGGACAGCAAGACACAGAGCCTGCTGATCGTCAACAACGCCACCAACGTGGTGATCAAGGTGTGCGAGTTTCAGTTCTGCAACGACCCCTTCCTGGACCACAAGAACAACAAGAGCTGGATGGAGAGCGAGTTCAGAGTGTATAGCAGCGCTAACAACTGCACCTTCGAGTACGTGAGCCAACCCTTCCTGATGGACCTGGAGGGCAAGCAAGGCAACTTCAAAAACCTGAGAGAGTTCGTGTTCAAGAACATCGACGGCTACTTCAAGATCTACAGCAAGCACACCCCCATCATCGTGAGAGAGCCCGAGGACCTGCCCCAAGGCTTCAGCGCCCTGGAGCCCCTGGTGGACCTGCCCATCGGCATCAACATCACAAGATTTCAGACCCTGCTGGCCCTGCACAGATCCTATCTGACCCCTGGGGACAGCAGCAG
CGGCTGGACCGCTGGCGCTGCCGCCTACTACGTGGGCTACCTGCAGCCTAGAACCTTCC
TGCTGAAGTACAACGAGAACGGCACAATCACCGATGCCGTCGACTGCGCCCTGGACCC
CCTGAGCGAGACCAAGTGCACCCTGAAGAGCTTCACCGTGGAGAAGGGCATCTATCAG
ACAAGCAACTTCAGAGTGCAGCCCACCGAGAGCATCGTGAGATTCCCCAACATCACCA
ACCTGTGCCCCTTCGACGAGGTGTTCAACGCCACAAGATTCGCTAGCGTGTACGCCTGG
AATAGAAAGAGAATCAGCAACTGCGTGGCCGACTACAGCGTGCTGTACAACCTGGCCC
CCTTCTTCACCTTCAAATGCTACGGCGTGAGCCCCACCAAGCTGAACGACCTGTGCTTC
ACCAACGTGTACGCCGACAGCTTCGTGATCAGAGGCGACGAGGTGAGACAGATCGCCC
CCGGGCAGACCGGCAACATCGCCGACTACAACTACAAGCTGCCCGACGACTTCACCGG
CTGCGTGATCGCCTGGAACAGCAACAAGCTCGACAGCAAGGTGAGCGGCAACTACAAC
TACCTGTACAGACTGTTCAGAAAGAGCAACCTGAAGCCCTTCGAGAGAGACATCAGCA
CCGAGATCTACCAAGCCGGCAACAAGCCCTGCAACGGCGTGGCCGGCTTCAACTGCTA
CTTCCCCCTGAGAAGCTACAGCTTCAGACCCACCTACGGCGTGGGCCATCAGCCCTACA
GAGTGGTCGTGCTGAGCTTCGAGCTGCTGCACGCCCCCGCCACCGTGTGCGGCCCCAA
GAAGAGCACCAACCTGGTGAAGAACAAGTGCGTGAACTTCAACTTCAACGGCCTGAAG
GGCACCGGCGTGCTGACCGAGAGCAACAAGAAGTTTCTGCCCTTTCAGCAGTTCGGCA
GAGACATCGCCGACACCACCGACGCCGTGAGAGACCCTCAGACCCTGGAGATCCTGGA
CATCACCCCCTGCTCCTTCGGCGGCGTGAGCGTGATCACCCCCGGCACCAACACAAGCA
ACCAAGTGGCCGTGCTGTACCAAGGCGTGAACTGCACCGAGGTGCCCGTGGCCATCCA
CGCCGATCAGCTGACCCCCACCTGGAGAGTGTACAGCACCGGCAGCAACGTGTTTCAG
ACAAGAGCCGGCTGCCTGATCGGCGCCGAGTACGTGAACAACAGCTACGAGTGCGACA
TCCCCATCGGCGCCGGCATCTGCGCTAGCTATCAGACACAGACCAAGAGCCACAGAAG
AGCTAGAAGCGTGGCTAGCCAAAGCATCATCGCCTACACCATGAGCCTGGGCGCCGAGA
ACAGCGTGGCCTACAGCAACAACAGCATCGCCATCCCCACCAACTTCACCATCAGCGTG
ACCACCGAAATTCTGCCCGTGAGCATGACCAAGACAAGCGTGGACTGCACCATGTACAT
CTGCGGCGACAGCACCGAGTGCAGCAACCTGCTCCTGCAGTACGGCAGCTTCTGCACA
CAGCTGAAGAGAGCCCTGACCGGCATCGCCGTGGAGCAAGACAAGAACACCCAAGAG
GTGTTCGCCCAAGTGAAGCAGATCTACAAGACCCCCCCCATCAAGTACTTCGGCGGCTT
CAACTTCAGCCAAATCCTGCCCGACCCTAGCAAGCCTAGCAAGAGAAGCTTCATCGAGG
ACCTGCTGTTCAACAAGGTGACCCTGGCCGACGCCGGCTTCATCAAGCAGTACGGCGA
CTGCCTGGGCGATATTGCCGCTAGAGACCTGATCTGCGCTCAGAAGTTCAAGGGCCTGA
CCGTGCTGCCCCCCCTGCTGACCGACGAGATGATCGCTCAGTATACAAGCGCCCTCCTC
GCTGGGACCATCACAAGCGGCTGGACATTCGGGGCCGGCGCTGCCCTGCAGATCCCCTT
CGCCATGCAGATGGCCTACAGATTCAACGGCATCGGCGTGACACAGAACGTGCTGTACG
AGAATCAGAAGCTGATCGCCAATCAGTTCAACAGCGCCATCGGCAAGATCCAAGACAG
CCTGAGCAGCACCGCTAGCGCCCTGGGCAAGCTGCAAGACGTGGTGAACCACAACGCC
CAAGCCCTGAACACCCTGGTGAAGCAGCTGAGCAGCAAGTTCGGCGCCATCAGCAGCG
TGCTGAACGACATCTTCAGCAGACTGGACCCCCCCGAGGCCGAGGTGCAGATCGACCG
GCTCATCACCGGCAGACTGCAGAGCCTGCAGACCTACGTGACACAGCAGCTGATCAGA
GCCGCCGAGATCAGAGCTAGCGCCAACCTGGCCGCCACCAAGATGAGCGAGTGCGTGC
TGGGGCAGAGCAAGAGAGTGGACTTCTGCGGCAAGGGCTACCACCTGATGAGCTTCCC
TCAGAGCGCCCCCCACGGCGTGGTGTTCCTGCACGTGACCTACGTGCCCGCCCAAGAG
AAGAACTTCACCACCGCTCCCGCCATTTGCCACGACGGCAAGGCCCACTTCCCTAGAGA
GGGCGTGTTCGTGAGCAACGGCACCCACTGGTTCGTGACACAGAGAAACTTCTACGAG
CCTCAGATCATCACCACCGACAACACCTTCGTCAGCGGCAACTGTGACGTGGTGATCGG
CATCGTGAACAATACCGTGTACGACCCCCTGCAGCCCGAACTGGACAGCTTCAAGGAG
GAGCTGGACAAGTACTTCAAGAACCACACAAGCCCCGACGTGGACCTGGGCGACATTA
GCGGCATCAACGCTAGCGTGGTGAACATTCAGAAGGAGATTGACAGACTGAACGAGGT
GGCCAAGAACCTGAACGAGAGCCTGATCGACCTGCAAGAGCTGGGCAAGTACGAGCA
GTACATCAAGTGGCCCTGGTACATCTGGCTGGGCTTCATCGCCGGCCTGATCGCCATCGT
GATGGTGACCATCATGCTGTGCTGCATGACAAGCTGCTGTAGCTGCCTGAAAGGCTGCT
GCAGCTGCGGCAGCTGCTGCAAGTTCGACGAGGACGACAGCGAGCCCGTGCTGAAGG
GCGTGAAGCTGCACTACACCTGA
SEQ ID No.12Ad-S-Omicron-BA.1疫苗S抗原的氨基酸序列
MFVFLVLLPLVSSQCVNLTTRTQLPPAYTNSFTRGVYYPDKVFRSSVLHSTQDLFLPFFSNVTWFHVISGTNGTKRFDNPVLPFNDGVYFASIEKSNIIRGWIFGTTLDSKTQSLLIVNNATNVVIKVCEFQFCNDPFLDHKNNKSWMESEFRVYSSANNCTFEYVSQPFLMDLEGKQGNFKNLREFVFKNIDGYFKIYSKHTPIIVREPEDLPQGFSALEPLVDLPIGINITRFQTLLALHRSYLTPGDSSSGWTAGAAAYYVGYLQPRTFLLKYNENGTITDAVDCALDPLSETKCTLKSFTVEKGIYQTSNFRVQPTESIVRFPNITNLCPFDEVFNATRFASVYAWNRKRISNCVADYSVLYNLAPFFTFKCYGVSPTKLNDLCFTNVYADSFVIRGDEVRQIAPGQTGNIADYNYKLPDDFTGCVIAWNSNKLDSKVSGNYNYLYRLFRKSNLKPFERDISTEIYQAGNKPCNGVAGFNCYFPLRSYSFRPTYGVGHQPYRVVVLSFELLHAPATVCGPKKSTNLVKNKCVNFNFNGLKGTGVLTESNKKFLPFQQFGRDIADTTDAVRDPQTLEILDITPCSFGGVSVITPGTNTSNQVAVLYQGVNCTEVPVAIHADQLTPTWRVYSTGSNVFQTRAGCLIGAEYVNNSYECDIPIGAGICASYQTQTKSHRRARSVASQSIIAYTMSLGAENSVAYSNNSIAIPTNFTISVTTEILPVSMTKTSVDCTMYICGDSTECSNLLLQYGSFCTQLKRALTGIAVEQDKNTQEVFAQVKQIYKTPPIKYFGGFNFSQILPDPSKPSKRSFIEDLLFNKVTLADAGFIKQYGDCLGDIAARDLICAQKFKGLTVLPPLLTDEMIAQYTSALLAGTITSGWTFGAGAALQIPFAMQMAYRFNGIGVTQNVLYENQKLIANQFNSAIGKIQDSLSSTASALGKLQDVVNHNAQALNTLVKQLSSKFGAISSVLNDIFSRLDPPEAEVQIDRLITGRLQSLQTYVTQQLIRAAEIRASANLAATKMSECVLGQSKRVDFCGKGYHLMSFPQSAPHGVVFLHVTYVPAQEKNFTTAPAICHDGKAHFPREGVFVSNGTHWFVTQRNFYEPQIITTDNTFVSGNCDVVIGIVNNTVYDPLQPELDSFKEELDKYFKNHTSPDVDLGDISGINASVVNIQKEIDRLNEVAKNLNESLIDLQELGKYEQYIKWPWYIWLGFIAGLIAIVMVTIMLCCMTSCCSCLKGCCSCGSCCKFDEDDSEPVLKGVKLHYT*
SEQ ID No.13Ad-S-Omicron-BA.2疫苗S抗原的核苷酸序列
ATGTTCGTGTTCCTGGTGCTGCTGCCCCTGGTGAGCAGCCAATGCGTGAACCTGATCACAAGAACACAGAGCTACACCAACAGCTTCACAAGAGGCGTGTACTACCCCGACAAGGTGTTCAGAAGCAGCGTCCTCCACAGCACCCAAGACCTGTTCCTGCCTTTCTTTAGCAACGTGACCTGGTTCCACGCCATCCACGTGAGCGGCACCAACGGCACCAAGAGATTCGACAACCCCGTGCTGCCCTTCAACGACGGCGTGTACTTCGCTAGCACCGAGAAGAGCAACATCATCAGAGGCTGGATCTTCGGCACCACCCTGGACAGCAAGACACAGAGCCTGCTGATCGTCAACAACGCCACCAACGTGGTGATCAAGGTGTGCGAGTTTCAGTTCTGCAACGACCCCTTCCTGGACGTGTACTACCACAAGAACAACAAGAGCTGGATGGAGAGCGAGTTCAGAGTGTACAGCAGCGCCAACAACTGCACCTTCGAGTACGTGAGCCAACCCTTCCTGATGGACCTGGAGGGCAAGCAAGGCAACTTCAAAAACCTGAGAGAGTTCGTGTTCAAGAACATCGACGGCTACTTCAAGATCTACAGCAAGCACACCCCCATCAACCTGGGCAGAGACCTGCCCCAAGGCTTCAGCGCCCTGGAGCCCCTGGTGGACCTGCCCATCGGCATCAACATCACAAGATTTCAGACCCTGCTGGCCCTGCACAGATCCTACCTCACACCTGGCGACAGCAGCTCCGGCTGGACCGCTGGCGCTGCCGCTTACTACGTGGGCTACCTGCAGCCTAGAACCTTCCTGCTGAAGTACAACGAGAACGGCACAATCACCGATGCCGTCGACTGCGCCCTGGACCCCCTGAGCGAGACCAAGTGCACCCTGAAGAGCTTCACCGTGGAGAAGGGCATCTATCAGACAAGCAACTTCAGAGTGCAGCCCACCGAGAGCATCGTGAGATTCCCCAACATCACCA
ACCTGTGCCCCTTCGACGAGGTGTTCAACGCCACAAGATTCGCTAGCGTGTACGCCTGG
AACAGAAAAAGAATCAGCAACTGCGTGGCCGACTACAGCGTGCTGTACAACTTCGCCC
CCTTCTTCGCCTTCAAATGCTACGGCGTGAGCCCCACCAAGCTGAACGACCTGTGCTTC
ACCAACGTGTACGCCGACAGCTTCGTGATCAGAGGCAACGAGGTGAGCCAAATCGCCC
CCGGGCAGACCGGCAACATCGCCGACTACAACTACAAGCTGCCCGACGACTTCACCGG
CTGCGTGATCGCCTGGAACTCCAACAAGCTCGACAGCAAGGTGGGCGGCAACTACAAC
TACCTGTACAGACTGTTCAGAAAGAGCAACCTGAAGCCCTTCGAGAGAGACATCAGCA
CCGAGATCTACCAAGCCGGCAACAAGCCCTGCAACGGCGTGGCCGGCTTCAACTGCTA
CTTCCCCCTGAGAAGCTACGGCTTCAGACCCACCTACGGCGTGGGCCATCAGCCCTACA
GAGTGGTCGTGCTGAGCTTCGAGCTGCTGCACGCCCCCGCCACCGTGTGCGGCCCCAA
GAAGAGCACCAACCTGGTGAAGAACAAGTGCGTGAACTTCAACTTCAACGGCCTGACC
GGCACCGGCGTGCTGACCGAGAGCAACAAGAAGTTCCTCCCCTTTCAGCAGTTCGGCA
GAGACATCGCCGACACCACCGACGCCGTGAGAGACCCTCAGACCCTGGAGATCCTGGA
CATCACCCCCTGCAGCTTCGGCGGCGTGAGCGTGATCACCCCCGGCACCAACACAAGC
AACCAAGTGGCCGTGCTGTACCAAGGCGTGAACTGCACCGAGGTGCCCGTGGCCATCC
ACGCCGATCAGCTGACCCCCACCTGGAGAGTGTACTCCACCGGCAGCAACGTGTTTCAG
ACAAGAGCCGGCTGCCTGATCGGCGCCGAGTACGTGAACAACAGCTACGAGTGCGACA
TCCCCATCGGCGCCGGCATCTGCGCTAGCTATCAGACACAGACCAAGAGCCACAGAAG
AGCTAGAAGCGTGGCTAGCCAAAGCATCATCGCCTACACCATGAGCCTGGGCGCCGAGA
ACAGCGTGGCCTACAGCAACAACAGCATCGCCATCCCCACCAACTTCACCATCAGCGTG
ACCACCGAGATCCTGCCTGTGAGCATGACCAAGACAAGCGTGGACTGCACCATGTACAT
CTGCGGCGACAGCACCGAGTGCAGCAACCTGCTCCTGCAGTACGGCAGCTTCTGCACA
CAGCTGAAGAGAGCCCTGACCGGCATCGCCGTGGAGCAAGACAAGAACACCCAAGAG
GTGTTCGCCCAAGTGAAGCAGATCTACAAGACCCCCCCCATCAAGTACTTCGGCGGCTT
CAACTTCAGCCAAATCCTGCCCGACCCTAGCAAGCCTAGCAAGCGGAGCTTCATCGAGG
ACCTGCTGTTCAACAAGGTGACCCTGGCCGACGCCGGCTTCATCAAGCAGTACGGCGA
CTGCCTCGGCGACATCGCCGCTAGAGACCTGATCTGCGCTCAGAAGTTCAATGGCCTGA
CCGTGCTGCCCCCCCTGCTGACCGACGAGATGATCGCTCAGTATACCTCCGCCCTGCTCG
CCGGGACCATCACAAGCGGCTGGACCTTCGGCGCCGGCGCCGCCCTCCAAATCCCCTTC
GCCATGCAGATGGCCTACAGATTCAACGGCATCGGCGTGACACAGAACGTGCTGTACGA
GAATCAGAAGCTGATCGCCAATCAGTTCAACAGCGCCATCGGCAAGATCCAAGACAGC
CTGAGCAGCACCGCTAGCGCCCTGGGCAAGCTGCAAGACGTGGTGAACCACAACGCCC
AAGCCCTGAACACCCTGGTGAAGCAGCTGAGCAGCAAGTTCGGCGCCATCAGCAGCGT
GCTGAACGATATCCTGAGCAGACTGGACCCCCCCGAGGCCGAGGTGCAGATCGATAGAC
TGATCACCGGCAGACTGCAGAGCCTGCAGACCTACGTGACACAGCAGCTGATCAGAGC
CGCCGAGATCAGAGCTAGCGCCAACCTGGCCGCCACCAAGATGAGCGAGTGCGTGCTG
GGGCAGAGCAAGAGAGTGGACTTCTGCGGCAAGGGCTACCACCTGATGAGCTTCCCTC
AGAGCGCCCCCCACGGCGTGGTGTTCCTGCACGTGACCTACGTGCCCGCCCAAGAGAA
GAACTTCACAACCGCCCCCGCCATCTGCCACGACGGCAAGGCCCACTTCCCTAGAGAG
GGCGTGTTCGTGAGCAACGGCACCCACTGGTTCGTGACACAGAGAAACTTCTACGAGC
CTCAGATCATCACCACCGACAACACCTTCGTGAGCGGCAACTGCGACGTGGTGATCGGG
ATCGTGAACAACACCGTGTACGATCCCCTGCAGCCCGAGCTGGACAGCTTCAAGGAGG
AGCTGGACAAGTACTTCAAGAACCACACAAGCCCCGACGTGGACCTGGGCGACATTAG
CGGCATCAACGCTAGCGTGGTGAACATTCAGAAGGAGATCGACAGACTGAATGAGGTG
GCCAAGAACCTGAACGAGAGCCTGATCGACCTGCAAGAGCTGGGCAAGTACGAGCAGT
ACATCAAGTGGCCCTGGTACATCTGGCTGGGCTTCATCGCCGGCCTGATCGCCATCGTGA
TGGTGACCATCATGCTGTGCTGCATGACAAGCTGCTGTAGCTGCCTGAAGGGCTGCTGC
AGCTGCGGCTCCTGCTGCAAGTTCGACGAGGACGACAGCGAGCCCGTGCTGAAGGGCG
TGAAGCTGCACTACACCTGA
SEQ ID No.14Ad-S-Omicron-BA.2疫苗S抗原的氨基酸序列
MFVFLVLLPLVSSQCVNLITRTQSYTNSFTRGVYYPDKVFRSSVLHSTQDLFLPFFSNVTWFHAIHVSGTNGTKRFDNPVLPFNDGVYFASTEKSNIIRGWIFGTTLDSKTQSLLIVNNATNVVIKVCEFQFCNDPFLDVYYHKNNKSWMESEFRVYSSANNCTFEYVSQPFLMDLEGKQGNFKNLREFVFKNIDGYFKIYSKHTPINLGRDLPQGFSALEPLVDLPIGINITRFQTLLALHRSYLTPGDSSSGWTAGAAAYYVGYLQPRTFLLKYNENGTITDAVDCALDPLSETKCTLKSFTVEKGIYQTSNFRVQPTESIVRFPNITNLCPFDEVFNATRFASVYAWNRKRISNCVADYSVLYNFAPFFAFKCYGVSPTKLNDLCFTNVYADSFVIRGNEVSQIAPGQTGNIADYNYKLPDDFTGCVIAWNSNKLDSKVGGNYNYLYRLFRKSNLKPFERDISTEIYQAGNKPCNGVAGFNCYFPLRSYGFRPTYGVGHQPYRVVVLSFELLHAPATVCGPKKSTNLVKNKCVNFNFNGLTGTGVLTESNKKFLPFQQFGRDIADTTDAVRDPQTLEILDITPCSFGGVSVITPGTNTSNQVAVLYQGVNCTEVPVAIHADQLTPTWRVYSTGSNVFQTRAGCLIGAEYVNNSYECDIPIGAGICASYQTQTKSHRRARSVASQSIIAYTMSLGAENSVAYSNNSIAIPTNFTISVTTEILPVSMTKTSVDCTMYICGDSTECSNLLLQYGSFCTQLKRALTGIAVEQDKNTQEVFAQVKQIYKTPPIKYFGGFNFSQILPDPSKPSKRSFIEDLLFNKVTLADAGFIKQYGDCLGDIAARDLICAQKFNGLTVLPPLLTDEMIAQYTSALLAGTITSGWTFGAGAALQIPFAMQMAYRFNGIGVTQNVLYENQKLIANQFNSAIGKIQDSLSSTASALGKLQDVVNHNAQALNTLVKQLSSKFGAISSVLNDILSRLDPPEAEVQIDRLITGRLQSLQTYVTQQLIRAAEIRASANLAATKMSECVLGQSKRVDFCGKGYHLMSFPQSAPHGVVFLHVTYVPAQEKNFTTAPAICHDGKAHFPREGVFVSNGTHWFVTQRNFYEPQIITTDNTFVSGNCDVVIGIVNNTVYDPLQPELDSFKEELDKYFKNHTSPDVDLGDISGINASVVNIQKEIDRLNEVAKNLNESLIDLQELGKYEQYIKWPWYIWLGFIAGLIAIVMVTIMLCCMTSCCSCLKGCCSCGSCCKFDEDDSEPVLKGVKLHYT*
SEQ ID No.15Ad-S-Omicron-BA.2.12.1疫苗S抗原的核苷酸序列
ATGTTCGTGTTCCTGGTGCTGCTGCCCCTGGTGAGCAGCCAATGCGTGAACCTGATCACAAGAACACAGAGCTACACCAACAGCTTCACAAGAGGCGTGTACTACCCCGACAAGGTGTTCAGAAGCAGCGTCCTCCACAGCACCCAAGACCTGTTCCTGCCTTTCTTTAGCAACGTGACCTGGTTCCACGCCATCCACGTGAGCGGCACCAACGGCACCAAGAGATTCGACAACCCCGTGCTGCCCTTCAACGACGGCGTGTACTTCGCTAGCACCGAGAAGAGCAACATCATCAGAGGCTGGATCTTCGGCACCACCCTGGACAGCAAGACACAGAGCCTGCTGATCGTCAACAACGCCACCAACGTGGTGATCAAGGTGTGCGAGTTTCAGTTCTGCAACGACCCCTTCCTGGACGTGTACTACCACAAGAACAACAAGAGCTGGATGGAGAGCGAGTTCAGAGTGTACAGCAGCGCCAACAACTGCACCTTCGAGTACGTGAGCCAACCCTTCCTGATGGACCTGGAGGGCAAGCAAGGCAACTTCAAAAACCTGAGAGAGTTCGTGTTCAAGAACATCGACGGCTACTTCAAGATCTACAGCAAGCACACCCCCATCAACCTGGGCAGAGACCTGCCCCAAGGCTTCAGCGCCCTGGAGCCCCTGGTGGACCTGCCCATCGGCATCAACATCACAAGATTTCAGACCCTGCTGGCCCTGCACAGATCCTACCTCACACCTGGCGACAGCAGCTCCGGCTGGACCGCTGGCGCTGCCGCTTACTACGTGGGCTACCTGCAGCCTAGAACCTTCCTGCTGAAGTACAACGAGAACGGCACAATCACCGATGCCGTCGACTGCGCCCTGGACCCCCTGAGCGAGACCAAGTGCACCCTGAAGAGCTTCACCGTGGAGAAGGGCATCTATCAGACAAGCAACTTCAGAGTGCAGCCCACCGAGAGCATCGTGAGATTCCCCAACATCACCAACCTGTGCCCCTTCGACGAGGTGTTCAACGCCACAAGATTCGCTAGCGTGTACGCCTGGAACAGAAAAAGAATCAGCAACTGCGTGGCCGACTACAGCGTGCTGTACAACTTCGCCCCCTTCTTCGCCTTCAAATGCTACGGCGTGAGCCCCACCAAGCTGAACGACCTGTGCTTCACCAACGTGTACGCCGACAGCTTCGTGATCAGAGGCAACGAGGTGAGCCAAATCGCCC
CCGGGCAGACCGGCAACATCGCCGACTACAACTACAAGCTGCCCGACGACTTCACCGG
CTGCGTGATCGCCTGGAACTCCAACAAGCTCGACAGCAAGGTGGGCGGCAACTACAAC
TATCAGTACAGACTGTTCAGAAAGAGCAACCTGAAGCCCTTCGAGAGAGACATCAGCA
CCGAGATCTACCAAGCCGGCAACAAGCCCTGCAACGGCGTGGCCGGCTTCAACTGCTA
CTTCCCCCTGAGAAGCTACGGCTTCAGACCCACCTACGGCGTGGGCCATCAGCCCTACA
GAGTGGTCGTGCTGAGCTTCGAGCTGCTGCACGCCCCCGCCACCGTGTGCGGCCCCAA
GAAGAGCACCAACCTGGTGAAGAACAAGTGCGTGAACTTCAACTTCAACGGCCTGACC
GGCACCGGCGTGCTGACCGAGAGCAACAAGAAGTTCCTCCCCTTTCAGCAGTTCGGCA
GAGACATCGCCGACACCACCGACGCCGTGAGAGACCCTCAGACCCTGGAGATCCTGGA
CATCACCCCCTGCAGCTTCGGCGGCGTGAGCGTGATCACCCCCGGCACCAACACAAGC
AACCAAGTGGCCGTGCTGTACCAAGGCGTGAACTGCACCGAGGTGCCCGTGGCCATCC
ACGCCGATCAGCTGACCCCCACCTGGAGAGTGTACTCCACCGGCAGCAACGTGTTTCAG
ACAAGAGCCGGCTGCCTGATCGGCGCCGAGTACGTGAACAACAGCTACGAGTGCGACA
TCCCCATCGGCGCCGGCATCTGCGCTAGCTATCAGACACAGACCAAGAGCCACAGAAG
AGCTAGAAGCGTGGCTAGCCAAAGCATCATCGCCTACACCATGAGCCTGGGCGCCGAGA
ACCTGGTGGCCTACAGCAACAACAGCATCGCCATCCCCACCAACTTCACCATCAGCGTG
ACCACCGAGATCCTGCCTGTGAGCATGACCAAGACAAGCGTGGACTGCACCATGTACAT
CTGCGGCGACAGCACCGAGTGCAGCAACCTGCTCCTGCAGTACGGCAGCTTCTGCACA
CAGCTGAAGAGAGCCCTGACCGGCATCGCCGTGGAGCAAGACAAGAACACCCAAGAG
GTGTTCGCCCAAGTGAAGCAGATCTACAAGACCCCCCCCATCAAGTACTTCGGCGGCTT
CAACTTCAGCCAAATCCTGCCCGACCCTAGCAAGCCTAGCAAGCGGAGCTTCATCGAGG
ACCTGCTGTTCAACAAGGTGACCCTGGCCGACGCCGGCTTCATCAAGCAGTACGGCGA
CTGCCTCGGCGACATCGCCGCTAGAGACCTGATCTGCGCTCAGAAGTTCAATGGCCTGA
CCGTGCTGCCCCCCCTGCTGACCGACGAGATGATCGCTCAGTATACCTCCGCCCTGCTCG
CCGGGACCATCACAAGCGGCTGGACCTTCGGCGCCGGCGCCGCCCTCCAAATCCCCTTC
GCCATGCAGATGGCCTACAGATTCAACGGCATCGGCGTGACACAGAACGTGCTGTACGA
GAATCAGAAGCTGATCGCCAATCAGTTCAACAGCGCCATCGGCAAGATCCAAGACAGC
CTGAGCAGCACCGCTAGCGCCCTGGGCAAGCTGCAAGACGTGGTGAACCACAACGCCC
AAGCCCTGAACACCCTGGTGAAGCAGCTGAGCAGCAAGTTCGGCGCCATCAGCAGCGT
GCTGAACGATATCCTGAGCAGACTGGACCCCCCCGAGGCCGAGGTGCAGATCGATAGAC
TGATCACCGGCAGACTGCAGAGCCTGCAGACCTACGTGACACAGCAGCTGATCAGAGC
CGCCGAGATCAGAGCTAGCGCCAACCTGGCCGCCACCAAGATGAGCGAGTGCGTGCTG
GGGCAGAGCAAGAGAGTGGACTTCTGCGGCAAGGGCTACCACCTGATGAGCTTCCCTC
AGAGCGCCCCCCACGGCGTGGTGTTCCTGCACGTGACCTACGTGCCCGCCCAAGAGAA
GAACTTCACAACCGCCCCCGCCATCTGCCACGACGGCAAGGCCCACTTCCCTAGAGAG
GGCGTGTTCGTGAGCAACGGCACCCACTGGTTCGTGACACAGAGAAACTTCTACGAGC
CTCAGATCATCACCACCGACAACACCTTCGTGAGCGGCAACTGCGACGTGGTGATCGGG
ATCGTGAACAACACCGTGTACGATCCCCTGCAGCCCGAGCTGGACAGCTTCAAGGAGG
AGCTGGACAAGTACTTCAAGAACCACACAAGCCCCGACGTGGACCTGGGCGACATTAG
CGGCATCAACGCTAGCGTGGTGAACATTCAGAAGGAGATCGACAGACTGAATGAGGTG
GCCAAGAACCTGAACGAGAGCCTGATCGACCTGCAAGAGCTGGGCAAGTACGAGCAGT
ACATCAAGTGGCCCTGGTACATCTGGCTGGGCTTCATCGCCGGCCTGATCGCCATCGTGA
TGGTGACCATCATGCTGTGCTGCATGACAAGCTGCTGTAGCTGCCTGAAGGGCTGCTGC
AGCTGCGGCTCCTGCTGCAAGTTCGACGAGGACGACAGCGAGCCCGTGCTGAAGGGCG
TGAAGCTGCACTACACCTGA
SEQ ID No.16Ad-S-Omicron-BA.2.12.1疫苗S抗原的氨基酸序列
MFVFLVLLPLVSSQCVNLITRTQSYTNSFTRGVYYPDKVFRSSVLHSTQDLFLPFFSNVTWFHAIHVSGTNGTKRFDNPVLPFNDGVYFASTEKSNIIRGWIFGTTLDSKTQSLLIVNNATNVVIKVCEFQFCNDPFLDVYYHKNNKSWMESEFRVYSSANNCTFEYVSQPFLMDLEGKQGNFKNLREFVFKNIDGYFKIYSKHTPINLGRDLPQGFSALEPLVDLPIGINITRFQTLLALHRSYLTPGDSSSGWTAGAAAYYVGYLQPRTFLLKYNENGTITDAVDCALDPLSETKCTLKSFTVEKGIYQTSNFRVQPTESIVRFPNITNLCPFDEVFNATRFASVYAWNRKRISNCVADYSVLYNFAPFFAFKCYGVSPTKLNDLCFTNVYADSFVIRGNEVSQIAPGQTGNIADYNYKLPDDFTGCVIAWNSNKLDSKVGGNYNYQYRLFRKSNLKPFERDISTEIYQAGNKPCNGVAGFNCYFPLRSYGFRPTYGVGHQPYRVVVLSFELLHAPATVCGPKKSTNLVKNKCVNFNFNGLTGTGVLTESNKKFLPFQQFGRDIADTTDAVRDPQTLEILDITPCSFGGVSVITPGTNTSNQVAVLYQGVNCTEVPVAIHADQLTPTWRVYSTGSNVFQTRAGCLIGAEYVNNSYECDIPIGAGICASYQTQTKSHRRARSVASQSIIAYTMSLGAENLVAYSNNSIAIPTNFTISVTTEILPVSMTKTSVDCTMYICGDSTECSNLLLQYGSFCTQLKRALTGIAVEQDKNTQEVFAQVKQIYKTPPIKYFGGFNFSQILPDPSKPSKRSFIEDLLFNKVTLADAGFIKQYGDCLGDIAARDLICAQKFNGLTVLPPLLTDEMIAQYTSALLAGTITSGWTFGAGAALQIPFAMQMAYRFNGIGVTQNVLYENQKLIANQFNSAIGKIQDSLSSTASALGKLQDVVNHNAQALNTLVKQLSSKFGAISSVLNDILSRLDPPEAEVQIDRLITGRLQSLQTYVTQQLIRAAEIRASANLAATKMSECVLGQSKRVDFCGKGYHLMSFPQSAPHGVVFLHVTYVPAQEKNFTTAPAICHDGKAHFPREGVFVSNGTHWFVTQRNFYEPQIITTDNTFVSGNCDVVIGIVNNTVYDPLQPELDSFKEELDKYFKNHTSPDVDLGDISGINASVVNIQKEIDRLNEVAKNLNESLIDLQELGKYEQYIKWPWYIWLGFIAGLIAIVMVTIMLCCMTSCCSCLKGCCSCGSCCKFDEDDSEPVLKGVKLHYT*
SEQ ID No.17Ad-S-Omicron-BA.4/5疫苗S抗原的核苷酸序列
ATGTTCGTGTTCCTGGTGCTGCTGCCCCTGGTGAGCAGCCAATGCGTGAACCTGATCACAAGAACACAGAGCTACACCAACAGCTTCACAAGAGGCGTGTACTACCCCGACAAGGTGTTCAGAAGCAGCGTCCTGCATTCCACCCAAGACCTGTTTCTCCCCTTCTTCAGCAACGTGACCTGGTTCCACGCCATCAGCGGCACCAACGGCACCAAGAGATTCGACAACCCCGTGCTGCCCTTCAACGACGGCGTGTACTTCGCTAGCACCGAGAAGAGCAACATCATCAGAGGCTGGATCTTCGGCACCACCCTGGACAGCAAGACACAGAGCCTGCTGATCGTGAACAACGCCACAAACGTGGTGATCAAGGTGTGCGAGTTTCAGTTCTGCAACGACCCCTTCCTGGACGTGTACTACCACAAGAACAACAAGAGCTGGATGGAGAGCGAGTTCCGGGTGTACAGCAGCGCCAACAACTGCACCTTCGAGTACGTGAGCCAACCCTTCCTGATGGACCTGGAGGGCAAGCAAGGCAACTTCAAGAACCTGAGAGAGTTCGTGTTCAAGAACATCGACGGCTACTTCAAGATCTACAGCAAGCACACCCCCATCAACCTGGGCAGAGACCTGCCCCAAGGCTTCAGCGCCCTGGAGCCCCTGGTGGACCTGCCCATCGGCATCAACATCACAAGATTTCAGACACTGCTCGCCCTGCACCGGAGCTACCTCACCCCTGGCGACAGCAGCTCCGGCTGGACCGCTGGGGCTGCCGCCTACTACGTGGGCTACCTGCAGCCTAGAACCTTCCTGCTGAAGTACAACGAGAACGGCACCATCACCGACGCTGTGGACTGTGCCCTGGACCCCCTGAGCGAGACCAAGTGCACCCTGAAGAGCTTCACCGTGGAGAAGGGCATCTATCAGACAAGCAACTTCAGAGTGCAGCCCACCGAGAGCATCGTGAGATTCCCCAACATCACCAACCTGTGCCCCTTCGACGAGGTGTTCAACGCCACAAGATTCGCTAGCGTGTACGCCTGGAACAGAAAGAGAATCAGCAACTGCGTGGCCGACTACAGCGTGCTGTACAACTTCGCCCCCTTCTTCGCCTTCAAATGCTACGGCGTCAGCCCCACCAAGCTGAACGACCTGTGCTTCACCAACGTGTACGCCGACAGCTTCGTGATCAGAGGCAACGAGGTGAGCCAAATCGCCCCCGGGCAGACCGGCAACATCGCCGACTATAACTACAAACTGCCCGACGACTTCACCGGCTGCGTGATCGCCTGGAACTCCAACAAGCTGGACAGCAAAGTGGGCGGCAACTACAACTACAGATACAGACTGTTCAGAAAGAGCAACCTGAAGCCCTTCGAGAGAGACATCAGCACCGAGATCTACCAAGCCGGCAACAAGCCCTGCAACGGCGTGGCCGGCGTGAACTGCTACTTCCCCC
TGCAGAGCTACGGCTTCAGACCCACCTACGGCGTGGGCCATCAGCCCTACAGAGTGGTC
GTGCTGAGCTTCGAGCTGCTGCACGCCCCTGCCACAGTGTGCGGCCCCAAGAAGAGCA
CCAACCTGGTGAAGAACAAGTGCGTGAACTTCAACTTCAACGGCCTGACCGGCACCGG
CGTGCTGACCGAGAGCAACAAGAAGTTCCTGCCCTTTCAGCAGTTCGGCAGAGACATC
GCCGACACCACCGATGCTGTGAGAGACCCTCAGACCCTGGAGATCCTGGACATCACCCC
TTGCAGCTTCGGCGGCGTGAGCGTGATCACCCCCGGCACCAACACAAGCAACCAAGTG
GCCGTGCTGTACCAAGGCGTCAACTGCACAGAGGTGCCCGTGGCCATCCACGCCGATCA
GCTGACCCCCACCTGGAGAGTGTACTCCACCGGCAGCAACGTGTTTCAGACAAGAGCC
GGCTGCCTGATCGGCGCCGAGTACGTGAACAACAGCTACGAGTGCGACATCCCCATCGG
CGCCGGCATCTGCGCTAGCTATCAGACACAGACCAAGAGCCACAGAAGAGCTAGAAGC
GTGGCTAGCCAAAGCATCATCGCCTACACCATGAGCCTGGGCGCCGAGAACAGCGTGG
CCTACAGCAACAACAGCATCGCCATCCCCACCAACTTCACCATCAGCGTGACCACCGAG
ATCCTGCCTGTGAGCATGACCAAGACAAGCGTGGACTGCACCATGTACATCTGCGGCGA
CAGCACCGAGTGCAGCAACCTGCTCCTGCAGTACGGCAGCTTCTGCACACAGCTGAAG
AGAGCCCTGACCGGCATCGCCGTGGAGCAAGACAAGAACACCCAAGAGGTGTTCGCCC
AAGTGAAGCAGATCTACAAGACCCCCCCCATCAAGTACTTCGGCGGCTTCAACTTCAGC
CAAATCCTGCCCGACCCTAGCAAGCCTAGCAAGCGGAGCTTCATCGAGGACCTGCTGTT
CAACAAGGTGACCCTGGCCGACGCCGGCTTCATCAAGCAGTACGGCGACTGCCTCGGC
GACATCGCTGCTAGAGACCTGATCTGCGCTCAGAAGTTCAACGGCCTCACAGTGCTGCC
CCCCCTGCTGACCGACGAGATGATCGCTCAGTACACCTCCGCTCTGCTGGCCGGCACAA
TTACATCCGGCTGGACCTTCGGCGCCGGCGCCGCCCTGCAAATCCCCTTCGCCATGCAG
ATGGCCTACAGATTCAACGGCATCGGCGTGACACAGAACGTGCTGTACGAGAATCAGAA
GCTGATCGCCAATCAGTTCAACAGCGCCATCGGCAAGATCCAAGACAGCCTGAGCAGC
ACCGCTAGCGCCCTGGGCAAGCTGCAAGACGTGGTGAACCACAACGCCCAAGCCCTGA
ACACCCTGGTGAAGCAGCTGAGCAGCAAGTTCGGCGCCATCTCCTCCGTCCTGAACGA
CATCCTGAGCAGACTGGACCCCCCCGAGGCCGAGGTGCAGATTGACAGACTGATTACCG
GCAGACTGCAGAGCCTGCAGACCTACGTGACACAGCAGCTGATCAGAGCCGCCGAGAT
CAGAGCTAGCGCCAACCTGGCCGCCACCAAGATGAGCGAGTGCGTGCTGGGGCAGAGC
AAGAGAGTGGACTTCTGCGGCAAGGGCTACCACCTGATGAGCTTCCCTCAGAGCGCCC
CCCACGGCGTGGTGTTCCTGCACGTGACCTACGTGCCCGCCCAAGAGAAGAACTTCAC
CACCGCTCCCGCCATCTGCCACGACGGCAAGGCCCACTTCCCTAGAGAGGGCGTGTTCG
TGAGCAACGGCACCCACTGGTTCGTGACACAGAGAAACTTCTACGAGCCTCAGATCATC
ACCACCGACAACACCTTCGTGAGCGGCAACTGCGACGTGGTGATCGGCATCGTCAACA
ACACCGTGTACGACCCCCTGCAACCCGAGCTGGACAGCTTCAAGGAGGAGCTGGACAA
GTACTTTAAGAACCACACAAGCCCCGACGTGGACCTGGGGGACATCTCCGGCATCAAC
GCTAGCGTGGTGAACATTCAGAAGGAAATTGACAGACTGAATGAGGTGGCCAAGAACC
TGAACGAGAGCCTGATCGACCTGCAAGAGCTGGGCAAGTACGAGCAGTACATCAAGTG
GCCCTGGTACATCTGGCTGGGCTTCATCGCCGGCCTGATCGCCATCGTGATGGTGACCAT
CATGCTGTGCTGCATGACAAGCTGCTGCAGCTGTCTGAAGGGCTGCTGCTCCTGCGGCA
GCTGCTGCAAGTTCGACGAGGACGACAGCGAGCCCGTGCTGAAGGGCGTGAAGCTGC
ACTACACCTGA
SEQ ID No.18Ad-S-Omicron-BA.4/5疫苗S抗原的氨基酸序列
MFVFLVLLPLVSSQCVNLITRTQSYTNSFTRGVYYPDKVFRSSVLHSTQDLFLPFFSNVTWFHAISGTNGTKRFDNPVLPFNDGVYFASTEKSNIIRGWIFGTTLDSKTQSLLIVNNATNVVIKVCEFQFCNDPFLDVYYHKNNKSWMESEFRVYSSANNCTFEYVSQPFLMDLEGKQGNFKNLREFVFKNIDGYFKIYSKHTPINLGRDLPQGFSALEPLVDLPIGINITRFQTLLALHRSYLTPGDSSSGWTAGAAAYYVGYLQPRTFLLKYNENGTITDAVDCALDPLSETKCTLKSFTVEKGIYQTSNFRVQPTESIVRFPNITNLCPFDEVFNATRFASVYAWNRKRISNCVADYSVLYNFAPFFAFKCYGVSPTKLNDLCFTNVYADSFVIRGNEVSQIAPGQTGNIADYNYKLPDDFTGCVIAWNSNKLDSKVGGNYNYRYRLFRKSNLKPFERDISTEIYQAGNKPCNGVAGVNCYFPLQSYGFRPTYGVGHQPYRVVVLSFELLHAPATVCGPKKSTNLVKNKCVNFNFNGLTGTGVLTESNKKFLPFQQFGRDIADTTDAVRDPQTLEILDITPCSFGGVSVITPGTNTSNQVAVLYQGVNCTEVPVAIHADQLTPTWRVYSTGSNVFQTRAGCLIGAEYVNNSYECDIPIGAGICASYQTQTKSHRRARSVASQSIIAYTMSLGAENSVAYSNNSIAIPTNFTISVTTEILPVSMTKTSVDCTMYICGDSTECSNLLLQYGSFCTQLKRALTGIAVEQDKNTQEVFAQVKQIYKTPPIKYFGGFNFSQILPDPSKPSKRSFIEDLLFNKVTLADAGFIKQYGDCLGDIAARDLICAQKFNGLTVLPPLLTDEMIAQYTSALLAGTITSGWTFGAGAALQIPFAMQMAYRFNGIGVTQNVLYENQKLIANQFNSAIGKIQDSLSSTASALGKLQDVVNHNAQALNTLVKQLSSKFGAISSVLNDILSRLDPPEAEVQIDRLITGRLQSLQTYVTQQLIRAAEIRASANLAATKMSECVLGQSKRVDFCGKGYHLMSFPQSAPHGVVFLHVTYVPAQEKNFTTAPAICHDGKAHFPREGVFVSNGTHWFVTQRNFYEPQIITTDNTFVSGNCDVVIGIVNNTVYDPLQPELDSFKEELDKYFKNHTSPDVDLGDISGINASVVNIQKEIDRLNEVAKNLNESLIDLQELGKYEQYIKWPWYIWLGFIAGLIAIVMVTIMLCCMTSCCSCLKGCCSCGSCCKFDEDDSEPVLKGVKLHYT*
有益效果：本发明将新冠病毒变异株的S蛋白突变体的986位赖氨酸(K)突变为脯氨酸(P)，987位缬氨酸(V)突变为脯氨酸(P)，以提高S蛋白稳定性以及表达水平。其中，Beta-Delta嵌合型S蛋白的基因序列是在Beta变异株的S蛋白突变位点的基础上加上Delta变异株S蛋白RBD的突变位点。再将S蛋白的氨基酸序列转换成核苷酸序列，并进行密码子优化，以获得编码SARS-CoV-2变异株S蛋白基因的多核苷酸。最后以E1/E3缺失的复制缺陷型人5型腺病毒为载体，制备抗SARS-CoV-2及变异株感染的疫苗，从而帮助宿主抵抗冠状病毒感染，特别是对突变病毒有较好的防治效果。
附图说明
图1为本发明实施例1新冠腺病毒疫苗S抗原结构示意图；
图2为试验1重组Ad5-Beta/Delta病毒全长基因构建示意图及重组腺病毒诱导刺突蛋白Spike的表达水平图；
图3为试验3重组腺病毒疫苗Ad5-WT、Ad5-Beta、Ad5-Delta、Ad5-Omicron BA.1、Ad5-Beta/Delta在血液中的抗RBD特异性结合抗体水平检测图；
图4分别为试验4重组腺病毒疫苗Ad5-WT、Ad5-Beta、Ad5-Delta、Ad5-OmicronBA.1、Ad5-Beta/Delta、Ad5-Omicron BA.4/5对假病毒的中和抗体检测结果图；
图5为试验5中重组腺病毒疫苗Ad5-Beta/Delta于肺泡灌洗液中抗RBD特异性IgG和IgA抗体的检测结果图；
图6为试验6腺病毒疫苗Ad5-Beta/Delta对组织驻留型T细胞的检测结果图；
图7为试验7中Ad5-Beta/Delta重组腺病毒疫苗对肺组织中抗原特异性T细胞的检测结果图；
图8为试验8中Ad5-Beta/Delta重组腺病毒疫苗生发中心反应的检测结果图。
具体实施方式
本发明首先提供了几种抗SARS-CoV-2及其变异株感染的疫苗，特别包含一种广谱的抗SARS-CoV-2感染的腺病毒疫苗。
所述的几种疫苗是含有经序列优化的SARS-CoV-2及Beta、Delta、Omicron变异株BA.1、BA.2、BA.2.12.1、BA.4/5和Beta-Delta变异株嵌合的S蛋白基因的重组E1/E3缺失的人5型、35型或26型或/和黑猩猩AdC68型和AdC7型的复制缺陷型腺病毒，SARS-CoV-2及其变异株S蛋白表达所用的信号肽是病毒自身的信号肽。
所述重组腺病毒疫苗被制备为肌肉注射剂、注射剂、滴鼻剂、喷雾剂或吸入剂。
在一个更为优选的实施方案中，所述重组腺病毒疫苗被制备为滴鼻剂、喷雾剂或吸入剂。
在一个优选实施方案中，为提高S蛋白的表达量和稳定性，Ad-S-Beta/Delta疫苗的目的基因不仅包括自身的信号肽，还包括在自身信号肽前额外添加的人tPA信号肽，而且除了将编码SARS-Cov-2的S蛋白氨基酸中的第986位K替换为P，第987位V替换为P；还对Furin蛋白酶切位点(RRAR)突变成GSAS。
本发明提供了制备上述的表达SARS-CoV-2及其变异株刺突蛋白的重组E1、E3联合缺失的人5型、35型或26型或/和黑猩猩AdC68型和AdC7型的复制缺陷型腺病毒的方法，所述方法包含以下步骤：
(1)构建包含编码SARS-CoV-2及Beta、Delta、Omicron BA.1、BA.2、BA.2.12.1、BA.4/5变异株和Beta-Delta变异株嵌合的S蛋白基因的穿梭质粒载体；
(2)将步骤(1)获得的穿梭质粒载体与骨架质粒共转293A宿主细胞，包装重组E1/E3联合缺失的人5型、35型或26型或/和黑猩猩AdC68型和AdC7型的复制缺陷型腺病毒；
(3)步骤(2)构建成功并鉴定正确的毒种在293细胞中逐级扩增，293细胞工厂或结合片状载体、微载体在生物反应器中扩增，或以悬浮细胞状态在生物反应器中规模化扩增；
(4)收集的培养物利用化学去垢剂裂解释放病毒，经澄清和超滤浓缩后，利用氯化铯超速离心或离子交换层析的的方法纯化，获得目的腺病毒。
上述步骤(4)中，离子交换层析的填料为Q Sepharose XL、Source 30Q或Source15Q等。
下面将结合实施例对本发明的方案进行解释。本领域技术人员将会理解，下面的实施例仅用于说明本发明，而不应视为限定本发明的范围。实施例中未注明具体技术或条件的，按照本领域内的文献所描述的技术或条件或者按照产品说明书进行。所用试剂或仪器未注明生产厂商者，均为可以通过市购获得的常规产品。
实施例1腺病毒疫苗的制备
1.S蛋白基因(野生型和各变异株)序列的优化和合成
选取病毒株(GenBank accession number:YP_009724390.1)为模板，获得野生型刺突(Spike,S)蛋白的基因序列，同时保留了S蛋白自身的信号肽。在此基础上，为保证蛋白提高表达水平的前提下，对密码子进行优化，随后优化后的序列进行基因合成。
在野生型病毒株(GenBank accession number:YP_009724390.1)刺突(Spike,S)蛋白序列的基础上，根据https://covariants.org/网站上提供的新冠病毒Beta、Delta、Omicron等各变异株的突变位点，获得各变异株S蛋白的核苷酸序列，同时保留了各S蛋白突变体自身的信号肽。Beta-Delta嵌合型S蛋白的基因序列是在Beta变异株的S蛋白突变位点的基础上加上Delta变异株S蛋白RBD的突变位点。在此基础上，为保证蛋白的稳定性及提高表达水平的前提下，对蛋白结构进行了突变。首先，将986位赖氨酸(L)替换为脯氨酸(P)，将987位缬氨酸(V)替换为脯氨酸(P)，以提高S蛋白稳定性以及表达水平。其次，对密码子进行优化。随后根据突变和优化的序列合成各变异株S蛋白基因。
按照以上方法合成的SARS-CoV-2及其变异株的S蛋白基因组有WT(野生型)、Beta、Delta、Beta/Delta嵌合型、Omicron BA.1、Omicron BA.4/5，其核苷酸序列分别如SEQ IDNo.3、SEQ ID No.5、SEQ ID No.7、SEQ ID No.9、SEQ ID No.11和SEQ ID No.17所示。
2.重组腺病毒新冠疫苗的包装
在基因合成过程中，将合成后的产物利用重组的克隆策略将S基因克隆至pDC316载体中，所得穿梭质粒(pDC316-S)。将以上构建好的包含野生型S和变异株S基因的pDC316-S分别与AdMax腺病毒系统的骨架质粒pBHGlox_E1,3Cre共转染HEK293细胞进行重组腺病毒的包装。过程如下：
1)将8×105个/孔HEK293A细胞接种于六孔板中，高糖DMEM+10％FBS培养基，置于37℃含5％CO2细胞培养箱中培养过夜。
2)第二天用高糖DMEM+2％FBS换液，将骨架质粒(pBHGlox_E1,3Cre)和穿梭质粒利用lipofectamine3000共转染HEK293A细胞。具体步骤为：每个转染孔取骨架质粒4μg，穿梭质粒2μg，用125μL Opti-MEM培养基进行稀释，然后加入12μL P3000试剂；另取一支1.5mlEP管，用125μL Opti-MEM培养基进行稀释7.5μL lipofectamine3000；将稀释的质粒和稀释的lipofectamine3000按1：1比例混合，室温孵育10-15分钟后加入细胞中，细胞继续培养，细胞长满后传代于25cm2细胞培养瓶中，每天观察细胞出毒迹象，待细胞长满瓶底时，再传入75cm2细胞培养瓶中，直到细胞出现明显噬斑，待细胞大部分病变并从底部脱落时进行收毒。
3)收集出毒的细胞培养物，1200rpm离心3分钟，吸取含病毒的上清，细胞沉淀用1/10培养体积的含病毒上清重悬，置于-80℃冰箱和37℃水浴锅中反复冻融三次。3000rpm离心20分钟，收集含病毒的上清液并与前述的含病毒上清合并，此即为腺病毒疫苗的毒种。
4)取50μL疫苗候选株毒种液，加入2μL蛋白酶K，50℃消化30min释放病毒基因组，以此为模版PCR扩增S基因序列，PCR产物电泳胶回收后测序鉴定，PCR扩增的条件如下：
变性：95℃10min
变性：95℃10s
退火：64℃30s
延伸：72℃2min
延伸：72℃5min
循环数40次
PCR扩增的引物如下：
pDC516-F1：ACACGTCAATGGGAAGTGAAA(SEQ ID No.19)
pDC516-R1：GCTAGACGATCCAGACATGAT(SEQ ID No.20)
3.重组腺病毒新冠疫苗的扩增
经鉴定正确的重组腺病毒疫苗毒种在293细胞中逐级扩增。具体过程如下：按MOI＝10加入80％-90％长满的293细胞，待大部分细胞病毒变圆后，收集病毒培养物，按照上述反复冻融的方法，制备主病毒种子库和工作病毒种子库。利用细胞工厂或生物反应器规模化扩增重组腺病毒疫苗，细胞大部分病变后收集病毒培养物。生物反应器扩增细胞和病毒的过程如下：首先在生物反应器中加入3-5g/LCytodex1微载体后灭菌，然后生物反应器中加入细胞培养液，当运行条件稳定在37℃，pH 7.0，DO 50％，50rpm时，消化收集细胞工厂扩增的HEK293细胞，接种于生物反应器内，接种细胞密度1.0-5.0×105个细胞/ml，补充细胞培养液至5L，反应器细胞培养的条件为温度37℃、转速为30-50rpm、pH7.15～7.25之间、DO30～50％，每天取样检测葡萄糖浓度、细胞密度和微载体上细胞的形态；当生物反应器内的细胞密度达到1.0～5.0×106个细胞/ml时，生物反应器接种重组腺病毒疫苗毒种，MOI为5-30，接种后每天取样检测葡萄糖浓度、培养上清和细胞沉淀中的病毒滴度、和观察微载体上细胞的形态；当大部分细胞重微载体上脱落时，终止培养，向生物反应器中加入病毒裂解液，终浓度为0.05％-1％ Tween 20 37℃裂解2-4小时，后收集病毒液。
4.重组腺病毒新冠疫苗的纯化
收集的病毒利用氯化铯超速离心或离子交换层析的的方法纯化，具体过程如下：
(1)氯化铯超速离心纯化腺病毒疫苗
收集的病毒培养物1200g离心10分钟，吸取含病毒的培养上清，细胞沉淀用1/10培养体积的含病毒上清重悬，-80℃冰箱和37℃水浴锅中反复冻融三次，3000rpm离心10-20分钟，吸取上清。含病毒的培养上清用100K-300K超滤膜包浓缩10倍；配制1.4g/ml氯化铯溶液(53g氯化铯+87ml 10mM Tris-HCl，PH 7.9)和1.2g/ml氯化铯溶液(氯化铯26.8+92ml10mMTris-HCl，PH 7.9)；在超速管中缓慢加入8ml 1.4g/ml氯化铯溶液，然后轻缓地加入6ml1.2g/ml氯化铯溶液，最后在不连续梯度顶部加入20ml含病毒上清液，配平后100000×g，4℃离心90分钟；离心结束后注射器抽吸出蓝色的病毒带，透析去除氯化铯后保存在-80℃。
(2)离子交换层析纯化腺病毒
收集病毒培养物，用0.05％-1％ Tween 20 37℃裂解2-4小时，裂解的培养物经1.2μm和0.45μm囊式滤器过滤澄清，使用100-300kD分子量的切向流膜包将样品浓缩10倍，随后用10体积的滤洗缓冲液(50mM Tris–HCl，2mM MgCl2，0–500mM NaCl，pH 8.0)进行滤洗，收集滤洗样品；滤洗的样品中加加入核酸酶，终浓度为10-50U/ml，37℃消化1-3小时后，样品利用Q Sepharose XL、Source 30Q或Source 15Q等填料进行阴离子交换层析，具体过程如下：平衡缓冲液20ml/min流速平衡5个柱体积，平衡结束后10ml/min上样，上样结束后平衡缓冲液平衡至电导水平；线性梯度洗脱样品，洗脱的条件为100％低盐缓冲液到100％高盐缓冲液，10V洗脱柱体积，流速为10ml/min，收集各个洗脱峰；洗脱结束后用2M NaCl缓冲液对柱子进行再生5～10个柱体积，速度为20ml/min。收集病毒峰，随后洗脱的病毒样品经透析或切向流过滤进行缓冲液置换(缓冲液成10mM Tris、10mM Na-PO4、150mM NaCl、2mMMgCl2、2％蔗糖、0.15％甘油、0.02％吐温80，pH7.6)。纯化后的腺病毒直接灌装，-20℃避光保存。
以下通过试验例证明本发明的有益效果。
其中，通过实施例1制备的腺病毒疫苗分别定义为Ad5-WT、Ad5-Beta、Ad5-Delta、Ad5-Omicron BA.1、Ad5-Omicron BA.4/5和Ad5-Beta/Delta重组腺病毒疫苗。
试验1重组腺病毒疫苗鉴定
我们首先通过人复制缺陷Ad5腺病毒载体分别表达SARS-CoV-2原始株、Beta株、Delta株、Omicron BA.1、Omicron BA.4/5株的全长刺突糖蛋白，同时我们设计和构建了在Beta毒株的全长基础上，在RBD区域增加Delta株的两个突变L452R和T478K的腺病毒Ad5-Beta/Delta(图2),通过免疫印迹Western blot的方法检测重组Ad5-Beta/Delta病毒感染293T细胞48小时后刺突蛋白Spike的表达水平。如图2所示，腺病毒Ad5-Beta/Delta感染293T细胞后可以诱导产生高水平的Spike蛋白表达，而空载腺病毒对照Ad5-Empty则不能，证明我们的重组腺病毒蛋白疫苗制备成功。
试验2动物准备及小鼠免疫和样品收集
6-8周龄的雌性BALB/c小鼠购买自维通利华公司，小鼠饲养在四川大学生物治疗国家重点实验室的无特殊病原体的环境中。首先将小鼠分为7组：(1)PBS组、(2)Ad5-Empty、(3)Ad5-WT、(4)Ad5-Beta、(5)Ad5-Delta、(6)Ad5-Omicron和(7)Ad5-Beta/Delta组，通过滴鼻免疫分别向BALB/c小鼠在第0，4，8周免疫以5×109病毒颗粒VP/只的剂量免疫小鼠。在第3、7周通过眼眶静脉采集血液样本，第11周处死小鼠并收集血液、肺泡灌洗液、肺和纵隔淋巴结组织，并将组织制成单细胞悬液进行流式检测。血液在4℃下以6000转/分离心10分钟。血清样品在使用前保存于-20℃，以便后续检测血清中的结合抗体和中和抗体。试验3抗RBD特异性抗体检测
用ELISA法检测血清中RBD特异性抗体。首先，我们用重组RBD蛋白(0.1μg/孔)包被96孔NUNC-MaxiSorp平板(Thermo Fisher Scientific,USA)，4℃，12小时。用1×PBST(1×PBS+0.1％ Tween-20)洗三次，用1％牛血清白蛋白(BSA)在37℃阻隔1小时。培养皿与2倍稀释梯度的血清样品在室温下孵育1小时，然后通过1×PBST洗涤三次。在培养皿中加入1:10000稀释的辣根过氧化物酶(HRP)-山羊抗小鼠IgG抗体，室温下孵育1小时，洗5次。在培养皿中加入3,3'，5,5'-四甲基联苯二胺(TMB)，暗处孵育10分钟。用1M H2SO4(100μl/孔)停止反应，在450nm处测定吸光度。
如图3所示，在接种后3、7、11周，重组腺病毒疫苗Ad5-WT、Ad5-Beta、Ad5-Delta、Ad5-Omicron.BA.1、Ad5-Beta/Delta都可以诱导产生高水平的血液中抗RBD特异性结合抗体，因此需要进一步检测血清中的中和抗体水平来评价各种重组腺病毒疫苗诱导的广谱中和免疫反应。
试验4假病毒中和试验
除了BA.3和BA.4/5假病毒从诺唯赞生物科技有限公司购买外，我们从吉满生物科技上海有限公司购买了SARS-CoV-2假病毒(GFP-Luciferase)的其他新冠病毒变异株假病毒。将在第11周处死小鼠后收集到的血清样本在60℃灭活30分钟后，用含有血清和抗生素的DMEM双有培养基三倍梯度稀释血清。稀释后的血清与荧光素酶假病毒(野生型WT，B.1.1.7,B.1.351,P.1,B.1.617,C.37,BA.1,BA.2,BA.2.12.1,BA.3,BA.4/5)在37℃孵育1小时。将1.2×104 293T/ACE2细胞添加到每个孔中表达报告基因。48小时后，去除感染细胞上清后，每孔加入100μl含荧光素酶底物的l裂解试剂。最后，用多模式微板阅读器(PerkinElmer,USA)检测。
如图4所示，Ad5-WT，Ad5-Beta,Ad5-Delta诱导出对自身病毒高水平的中和抗体，但由于Omicron毒株的强的免疫逃逸能力，这三种疫苗诱导针对Omicron亚变的中和抗体水平较低。Ad5-Omicron-BA.1/BA.5疫苗可以诱导出针对Omicron亚变体的高水平中和抗体，但是其无法诱导出针对WT、Alpha、Beta、Delta等高水平的中和抗体。与其他腺病毒疫苗相比，Ad5-Beta/Delta可以诱导出针对包括WT、Alpha、Beta、Delta、Omicron BA.1、BA.2、BA.2.12.1、BA.3及BA.4/5的高水平广谱性中和抗体。
试验5肺泡灌洗液中抗RBD特异性IgG和IgA抗体的检测
用ELISA法检测肺泡灌洗液中的RBD特异性抗体IgG和IgA。首先，我们用重组RBD蛋白(0.1μg/孔)包被96孔NUNC-MaxiSorp平板(Thermo Fisher Scientific,USA)，4℃，12小时。用1×PBST(1×PBS+0.1％ Tween-20)洗三次，用1％牛血清白蛋白(BSA)在37℃阻隔1小时。培养皿与2倍稀释梯度的肺泡灌洗液样品在室温下孵育1小时，然后通过1×PBST洗涤三次。在培养皿中加入1:10000稀释的辣根过氧化物酶(HRP)-山羊抗小鼠IgG或IgA(1：5000稀释)抗体，室温下孵育1小时，洗5次。在培养皿中加入3,3'，5,5'-四甲基联苯二胺(TMB)，暗处孵育10分钟。用1M H2SO4(100μl/孔)停止反应，在450nm处测定吸光度。
如图5所示，在接种后11周，重组腺病毒疫苗Ad5-Beta/Delta诱导产生高水平的肺泡灌洗液中抗RBD特异性结合抗体IgG和IgA，这说明我们的重组腺病毒疫苗可以在呼吸道黏膜部分产生强烈的抗体保护。
试验6肺泡灌洗液中组织驻留型T细胞的检测
使用流式细胞术检测肺泡灌洗液中组织驻留型T细胞(Tissue resident Tcells，TRM)的数量。将收集好的1ml肺泡灌洗液离心(400×g，5分钟)，弃上清后获得细胞沉淀，使用100μlPBS缓冲液重悬细胞后，加入PerCP/Cyanine5.5-conjugated anti-mouseCD3(BioLegend,100718),Brilliant Violet 421-conjugated anti-mouse CD4(BioLegend,100412),Brilliant Violet510-conjugated anti-mouse CD8,PE-conjugated anti-mouse CD44,FITC-conjugated anti-mouse CD69,and APC-conjugatedanti-mouse CD103抗体后，在4℃孵育30分钟。用PBS洗涤一次后，重选细胞，并通过流式细胞术检测。
如图6所示，滴鼻免疫Ad5-Beta/Delta后，相比较于PBS处理的小鼠，腺病毒疫苗可以在肺组织募集大量的组织驻留型CD4+和CD8+T细胞，产生细胞免疫保护。
试验7肺组织中抗原特异性T细胞的检测
使用流式细胞术检测肺组织中抗原特异性T细胞的检测。将收集的肺组织剪碎至1mm3大小后，加入配置好的胶原酶消化液，并在37℃孵育1小时，将组织消化液过70uM筛网得到单细胞悬液。肺组织单细胞在含有刺突蛋白Spike肽库的双有1640培养基中刺激并培养12h，进行细胞内细胞因子染色(ICS)。在1640培养基中加入10％胎牛血清、100μg/ml链霉素、100U/ml青霉素、1mM丙酮酸(均来自美国Gibco公司)、50μMβ-巯基乙醇和20U/ml IL-2(均来自Sigma-Aldrich公司)。为了阻止细胞内细胞因子的分泌，brefeldin A(BFA,BDBiosciences)在染色前孵育6小时。细胞在1×PBS冷液中洗涤，在4℃下用PerCP/Cyanine5.5-conjugated anti-mouse CD3(BioLegend,100718),APC-conjugated anti-mouse CD4(BioLegend,100412),FITC-conjugated anti-mouse CD8,PE-conjugatedanti-mouse CD44抗体染色30分钟。然后将细胞固定并打孔，以促进细胞内用PE-Cy7-conjugated anti-mouse IFN-γ,Brilliant Violet510-conjugated anti-mouse TNF-α(所有流式抗体来自BioLegend)在室温下放置2小时染色。用1×PBS洗细胞，再用流式细胞术检测。
T细胞反应是临床结果的主要决定因素，对预防SARS-CoV-2具有重要作用。如图7所示，滴鼻免疫Ad5-Beta/Delta疫苗后诱导产生强大的抗原特异性细胞免疫，包括分泌IFN-γ和TNF-α的T细胞。
试验8生发中心反应的检测
纵隔淋巴细胞单细胞悬液制备完成后，我们用PerCP/Cyanine5.5-conjugatedanti-mouse CD3,PE-Cy7-conjugated anti-mouse CD45R/B220,Brilliant Violet 421-conjugated anti-mouse GL-7,APC-conjugated anti-mouse CD95,Brilliant Violet421-conjugated anti-mouse CD19,APC-conjugated anti-mouse CD4,PE-conjugatedanti-mouse CXCR5,and Brilliant Violet510-conjugated PD-1抗体对Tfh细胞、GC B细胞进行染色(流式抗体均来自BioLegend)。细胞在黑暗中4℃染色。30分钟后，用1×PBS冲洗细胞，用流式细胞仪检测。
持久的记忆反应在再次感染时提供快速有效的保护性免疫。Tfh和GCB细胞的升高代表疫苗可以对机体提供长效的保护性免疫。如图8所示，滴鼻免疫Ad5-Beta/Delta疫苗后可以诱导纵隔淋巴结内Tfh和GCB细胞明显升高，证明我们的重组腺病毒疫苗可以提高长效的免疫保护。
以上仅是本发明的优选实施方案，应当指出的是，上述优选实施方式不应视为对本发明的限制，本发明的保护范围应当以权利要求所限定的范围为准，对于本技术领域的普通技术人员来说，在不脱离本发明的精神和范围内，还可以做出若干改进和润饰，这些改进和润饰也应视为本发明的保护范围。
Claims (15)
1.编码SARS-CoV-2或变异株S蛋白基因的多核苷酸，其特征在于：所述的多核苷酸序列如SEQ ID No.3所示；或者在SEQ ID No.3核苷酸序列上经取代和/或缺失和/或插入至少一个核苷酸，且与SEQ ID No.3具有相同或相似的生物学活性的核苷酸序列。
2.如权利要求1所述的多核苷酸，其特征在于：所述多核苷酸序列包括S蛋白或其突变体自身的信号肽，所述自身的信号肽核苷酸序列如SEQ ID No.1所示；优选地，所述多核苷酸序列包括S蛋白或其突变体自身的信号肽和在自身信号肽前额外添加的人tPA信号肽。
3.如权利要求1所述的多核苷酸，其特征在于：所述的多核苷酸序列编码的SARS-CoV-2变异株S蛋白是将编码SARS-CoV-2的S蛋白氨基酸中的第986位K替换为P，第987位V替换为P；所述编码SARS-CoV-2的S蛋白氨基酸序列如SEQ ID No.4所示；优选地，所述的多核苷酸序列编码的SARS-CoV-2变异株S蛋白还包括将Furin蛋白酶切位点RRAR突变成GSAS。
4.如权利要求3所述的多核苷酸，其特征在于：所述多核苷酸序列选自新冠病毒野生型、Alpha、Beta、Gamma、Delta、Omicron毒株中的S蛋白核苷酸序列中的至少一种。
5.如权利要求4所述的多核苷酸，其特征在于：所述多核苷酸序列选自SEQ ID No.3、SEQ ID No.5、SEQ ID No.7、SEQ ID No.9、SEQ ID No.11、SEQ ID No.13、SEQ ID No.15或SEQ ID No.17中的至少一种；优选地，所述多核苷酸序列选自SEQ ID No.9、SEQ ID No.11、SEQ ID No.13、SEQ ID No.15或SEQ ID No.17中的至少一种。
6.如权利要求4所述的多核苷酸，其特征在于：所述的多核苷酸序列编码的SARS-Cov-2及变异株S蛋白的氨基酸序列选自SEQ ID No.4、SEQ ID No.6、SEQ ID No.8、SEQ IDNo.10、SEQ ID No.12、SEQ ID No.14、SEQ ID No.16或SEQ ID No.18中的至少一种；优选地，所述的多核苷酸序列编码的SARS-Cov-2变异株S蛋白的氨基酸序列选自SEQ ID No.10、SEQ IDNo.12、SEQ ID No.14、SEQ ID No.16或SEQ ID No.18中的至少一种。
7.如权利要求3所述的多核苷酸，其特征在于：所述的多核苷酸序列编码的SARS-CoV-2变异株S蛋白的氨基酸序列是在Beta变异株的S蛋白突变位点的基础上加上Delta变异株S蛋白RBD的突变位点。
8.能够表达权利要求1-7任一项所述多核苷酸的复制缺陷型重组腺病毒载体。
9.如权利要求8所述的重组腺病毒载体，其特征在于：所述复制缺陷型重组腺病毒载体选自人5型、35型或26型或/和黑猩猩AdC68型或AdC7型的复制缺陷型腺病毒；优选地，选自E1、E3联合缺失的人5型复制缺陷型腺病毒。
10.预防和/或治疗SARS-CoV-2或其变异株感染的疫苗，其特征在于：其含有能够表达权利要求1-7任一项所述的多核苷酸；或含有权利要求8或9所述的复制缺陷型重组腺病毒载体。
11.如权利要求10所述的疫苗，其特征在于：所述疫苗还包括药学上可接受的佐剂、载体、稀释剂或赋形剂。
12.如权利要求10所述的疫苗，其特征在于：所述疫苗剂型为肌肉注射剂、注射剂、滴鼻剂、喷雾剂或吸入剂；优选地，所述疫苗为滴鼻剂、喷雾剂或吸入剂。
13.制备权利要求8或9中的腺病毒的方法，其特征在于：包括以下步骤：构建含有权利要求1-7任一项所述多核苷酸的穿梭质粒载体；然后将构建的穿梭质粒载体与骨架质粒一起转染入宿主细胞，培养宿主细胞；获得复制缺陷重组腺病毒，然后扩大培养、纯化，即可。
14.如权利要求13所述的方法，其特征在于：满足以下至少一项：
所述穿梭质粒载体为pDC316-S、pDC315-S、pDC516-S或pDC515-S中至少一种；
所述骨架质粒为pBHGlox_E1,3Cre、pBHGlox_E1,3FLP中至少一种；
所述宿主细胞为HEK293。
15.权利要求1-7任一项所述的多核苷酸，权利要求10-12任一项所述的疫苗在制备预防和/或治疗呼吸道病毒感染或致病药物中的用途；优选地，所述呼吸道病毒感染是SARS-CoV-2或其变异株感染引起的。
Applications Claiming Priority (2)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN202210802239 | 2022-07-07 | ||
| CN2022108022395 | 2022-07-07 |
Publications (1)
| Publication Number | Publication Date |
|---|---|
| CN116970624A true CN116970624A (zh) | 2023-10-31 |
Family
ID=88472063
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN202310726118.1A Pending CN116970624A (zh) | 2022-07-07 | 2023-06-19 | 抗SARS-CoV-2及变异株感染的腺病毒疫苗 |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN116970624A (zh) |
-
2023
- 2023-06-19 CN CN202310726118.1A patent/CN116970624A/zh active Pending
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| CN112076315B (zh) | 新冠病毒s蛋白和铁蛋白亚基融合的纳米抗原颗粒、新冠疫苗及其制备方法和应用 | |
| CN111218459B (zh) | 一种以人复制缺陷腺病毒为载体的重组新型冠状病毒疫苗 | |
| CN112618707B (zh) | 一种SARS-CoV-2冠状病毒疫苗及其制备方法 | |
| KR102044051B1 (ko) | 터미널 말단이 변경된 재조합 아데노바이러스의 배치 | |
| EP3184119A1 (en) | Chromatography based purification strategies for measles scaffold based viruses | |
| CN110156896B (zh) | 重组口蹄疫病毒样颗粒及其制备方法和应用 | |
| US11931410B1 (en) | SARS-CoV-2 mRNA vaccine and preparation method and use thereof | |
| WO2024008014A1 (zh) | 抗SARS-CoV-2或其突变体感染的药物组合物及其联合用药物 | |
| CN105274142B (zh) | 复制型重组人55型腺病毒载体及其制备方法和应用 | |
| CN117551677A (zh) | 一种以5型腺病毒为载体的猴痘病毒特异性融合蛋白疫苗 | |
| CN113817753B (zh) | 表达SARS-CoV-2纤突蛋白或其变异体SΔ21的假型化VSV病毒构建和应用 | |
| Li et al. | Development and characterization of Rift Valley fever virus-like particles | |
| CN116970624A (zh) | 抗SARS-CoV-2及变异株感染的腺病毒疫苗 | |
| CN113248577B (zh) | 一种以腺病毒为载体的冠状病毒疫苗及其制备方法 | |
| CN114657154B (zh) | 一种羊传染性脓疱病毒减毒株的制备方法及其应用 | |
| EP4014991A1 (en) | Modified parapoxvirus having increased immunogenicity | |
| CN111518815B (zh) | 通用埃博拉病毒病免疫球蛋白及其制备方法与应用 | |
| CN116549627A (zh) | 基于腺病毒载体的广谱新冠疫苗及其应用 | |
| CN115094088A (zh) | 一种pSFV-flagX-CMV复制子质粒、制备方法和鸡尾酒混合疫苗 | |
| KR20230008707A (ko) | 코로나바이러스 치료용 백신 조성물 | |
| CN112390860A (zh) | Eb病毒抗原表位及其应用 | |
| JP2002017370A (ja) | 遺伝子組換えワクシニアウイルスワクチン | |
| WO2024082795A1 (zh) | 抗SARS-CoV-2奥密克戎突变株XBB及其亚型感染的蛋白及疫苗 | |
| WO2024193380A1 (zh) | 抗呼吸道合胞病毒感染的疫苗 | |
| KR102277089B1 (ko) | A형 간염 바이러스의 제조방법 및 상기의 방법에 따라 제조된 a형 간염 바이러스 |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| PB01 | Publication | ||
| PB01 | Publication | ||
| SE01 | Entry into force of request for substantive examination | ||
| SE01 | Entry into force of request for substantive examination |