CN1632124A - 编码h5亚型禽流感病毒血凝素蛋白的基因及其应用 - Google Patents
编码h5亚型禽流感病毒血凝素蛋白的基因及其应用 Download PDFInfo
- Publication number
- CN1632124A CN1632124A CN 200410009866 CN200410009866A CN1632124A CN 1632124 A CN1632124 A CN 1632124A CN 200410009866 CN200410009866 CN 200410009866 CN 200410009866 A CN200410009866 A CN 200410009866A CN 1632124 A CN1632124 A CN 1632124A
- Authority
- CN
- China
- Prior art keywords
- gene
- dna
- asn
- fragment
- leu
- 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.)
- Granted
Links
- 108090000623 proteins and genes Proteins 0.000 title claims abstract description 103
- 241000712461 unidentified influenza virus Species 0.000 title claims abstract description 38
- 101710154606 Hemagglutinin Proteins 0.000 title claims abstract description 37
- 101710093908 Outer capsid protein VP4 Proteins 0.000 title claims abstract description 37
- 101710135467 Outer capsid protein sigma-1 Proteins 0.000 title claims abstract description 37
- 101710176177 Protein A56 Proteins 0.000 title claims abstract description 37
- 230000002163 immunogen Effects 0.000 claims abstract description 24
- 239000002773 nucleotide Substances 0.000 claims abstract description 10
- 125000003729 nucleotide group Chemical group 0.000 claims abstract description 10
- 210000004027 cell Anatomy 0.000 claims description 54
- 108010041986 DNA Vaccines Proteins 0.000 claims description 16
- 229940021995 DNA vaccine Drugs 0.000 claims description 16
- 239000013604 expression vector Substances 0.000 claims description 8
- 238000002360 preparation method Methods 0.000 claims description 7
- 210000003292 kidney cell Anatomy 0.000 claims description 4
- 108010008038 Synthetic Vaccines Proteins 0.000 claims description 3
- 241000700662 Fowlpox virus Species 0.000 claims description 2
- 210000003527 eukaryotic cell Anatomy 0.000 claims description 2
- 210000002950 fibroblast Anatomy 0.000 claims description 2
- 229940124551 recombinant vaccine Drugs 0.000 claims description 2
- 229940031626 subunit vaccine Drugs 0.000 claims description 2
- 210000003837 chick embryo Anatomy 0.000 claims 1
- 241000287828 Gallus gallus Species 0.000 abstract description 49
- 108020004705 Codon Proteins 0.000 abstract description 27
- 241000272814 Anser sp. Species 0.000 abstract description 14
- 150000001413 amino acids Chemical class 0.000 abstract description 14
- 239000000185 hemagglutinin Substances 0.000 abstract description 14
- 102000004169 proteins and genes Human genes 0.000 abstract description 13
- 230000001717 pathogenic effect Effects 0.000 abstract description 12
- 229960005486 vaccine Drugs 0.000 abstract description 11
- 229940033326 influenza DNA vaccine Drugs 0.000 abstract description 9
- 238000010353 genetic engineering Methods 0.000 abstract description 6
- 230000036961 partial effect Effects 0.000 abstract description 4
- 230000010415 tropism Effects 0.000 abstract description 4
- 239000012634 fragment Substances 0.000 description 134
- 108020004414 DNA Proteins 0.000 description 72
- 239000013612 plasmid Substances 0.000 description 55
- 235000013330 chicken meat Nutrition 0.000 description 48
- 239000000047 product Substances 0.000 description 39
- 230000003053 immunization Effects 0.000 description 31
- 108091034117 Oligonucleotide Proteins 0.000 description 30
- 238000002649 immunization Methods 0.000 description 30
- 206010064097 avian influenza Diseases 0.000 description 24
- 208000002979 Influenza in Birds Diseases 0.000 description 23
- 101150039660 HA gene Proteins 0.000 description 20
- 230000014509 gene expression Effects 0.000 description 20
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 description 18
- 238000006243 chemical reaction Methods 0.000 description 18
- 238000000034 method Methods 0.000 description 17
- 230000009182 swimming Effects 0.000 description 16
- 238000001962 electrophoresis Methods 0.000 description 15
- 241000700605 Viruses Species 0.000 description 14
- 238000001514 detection method Methods 0.000 description 13
- 239000000499 gel Substances 0.000 description 13
- 238000001890 transfection Methods 0.000 description 13
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Chemical compound O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 description 13
- 108700010070 Codon Usage Proteins 0.000 description 12
- 230000002441 reversible effect Effects 0.000 description 12
- 238000005516 engineering process Methods 0.000 description 11
- 235000018102 proteins Nutrition 0.000 description 11
- 239000000243 solution Substances 0.000 description 11
- 229940024606 amino acid Drugs 0.000 description 10
- 235000001014 amino acid Nutrition 0.000 description 10
- 239000012528 membrane Substances 0.000 description 10
- 238000005457 optimization Methods 0.000 description 10
- 238000010166 immunofluorescence Methods 0.000 description 9
- 238000012163 sequencing technique Methods 0.000 description 9
- 238000012360 testing method Methods 0.000 description 9
- 239000000020 Nitrocellulose Substances 0.000 description 8
- 239000008367 deionised water Substances 0.000 description 8
- 229910021641 deionized water Inorganic materials 0.000 description 8
- 238000007857 nested PCR Methods 0.000 description 8
- 229920001220 nitrocellulos Polymers 0.000 description 8
- 108010006785 Taq Polymerase Proteins 0.000 description 7
- 108091007433 antigens Proteins 0.000 description 7
- 239000000203 mixture Substances 0.000 description 7
- 238000011144 upstream manufacturing Methods 0.000 description 7
- 238000001262 western blot Methods 0.000 description 7
- 102000004190 Enzymes Human genes 0.000 description 6
- 108090000790 Enzymes Proteins 0.000 description 6
- OKKJLVBELUTLKV-UHFFFAOYSA-N Methanol Chemical compound OC OKKJLVBELUTLKV-UHFFFAOYSA-N 0.000 description 6
- 102000036639 antigens Human genes 0.000 description 6
- 230000001681 protective effect Effects 0.000 description 6
- 238000011160 research Methods 0.000 description 6
- 210000002966 serum Anatomy 0.000 description 6
- 238000012546 transfer Methods 0.000 description 6
- OJHZNMVJJKMFGX-RNWHKREASA-N (4r,4ar,7ar,12bs)-9-methoxy-3-methyl-1,2,4,4a,5,6,7a,13-octahydro-4,12-methanobenzofuro[3,2-e]isoquinoline-7-one;2,3-dihydroxybutanedioic acid Chemical compound OC(=O)C(O)C(O)C(O)=O.O=C([C@@H]1O2)CC[C@H]3[C@]4([H])N(C)CC[C@]13C1=C2C(OC)=CC=C1C4 OJHZNMVJJKMFGX-RNWHKREASA-N 0.000 description 5
- DHMQDGOQFOQNFH-UHFFFAOYSA-N Glycine Chemical compound NCC(O)=O DHMQDGOQFOQNFH-UHFFFAOYSA-N 0.000 description 5
- 238000004458 analytical method Methods 0.000 description 5
- 239000000427 antigen Substances 0.000 description 5
- 239000000872 buffer Substances 0.000 description 5
- 210000003555 cloaca Anatomy 0.000 description 5
- 230000034994 death Effects 0.000 description 5
- 231100000517 death Toxicity 0.000 description 5
- 238000011161 development Methods 0.000 description 5
- 230000029087 digestion Effects 0.000 description 5
- 238000002955 isolation Methods 0.000 description 5
- 239000013598 vector Substances 0.000 description 5
- ZHNUHDYFZUAESO-UHFFFAOYSA-N Formamide Chemical compound NC=O ZHNUHDYFZUAESO-UHFFFAOYSA-N 0.000 description 4
- 241000725303 Human immunodeficiency virus Species 0.000 description 4
- KDXKERNSBIXSRK-UHFFFAOYSA-N Lysine Natural products NCCCCC(N)C(O)=O KDXKERNSBIXSRK-UHFFFAOYSA-N 0.000 description 4
- 241001465754 Metazoa Species 0.000 description 4
- YBAFDPFAUTYYRW-UHFFFAOYSA-N N-L-alpha-glutamyl-L-leucine Natural products CC(C)CC(C(O)=O)NC(=O)C(N)CCC(O)=O YBAFDPFAUTYYRW-UHFFFAOYSA-N 0.000 description 4
- 241000283973 Oryctolagus cuniculus Species 0.000 description 4
- FAPWRFPIFSIZLT-UHFFFAOYSA-M Sodium chloride Chemical compound [Na+].[Cl-] FAPWRFPIFSIZLT-UHFFFAOYSA-M 0.000 description 4
- 239000011543 agarose gel Substances 0.000 description 4
- 230000003321 amplification Effects 0.000 description 4
- 230000015572 biosynthetic process Effects 0.000 description 4
- 230000000295 complement effect Effects 0.000 description 4
- 230000000694 effects Effects 0.000 description 4
- 210000002257 embryonic structure Anatomy 0.000 description 4
- 230000028993 immune response Effects 0.000 description 4
- 238000000338 in vitro Methods 0.000 description 4
- 210000004962 mammalian cell Anatomy 0.000 description 4
- 238000003199 nucleic acid amplification method Methods 0.000 description 4
- 239000002244 precipitate Substances 0.000 description 4
- 238000002415 sodium dodecyl sulfate polyacrylamide gel electrophoresis Methods 0.000 description 4
- 238000003786 synthesis reaction Methods 0.000 description 4
- 230000014616 translation Effects 0.000 description 4
- QTBSBXVTEAMEQO-UHFFFAOYSA-N Acetic acid Chemical compound CC(O)=O QTBSBXVTEAMEQO-UHFFFAOYSA-N 0.000 description 3
- 239000006144 Dulbecco’s modified Eagle's medium Substances 0.000 description 3
- KCXVZYZYPLLWCC-UHFFFAOYSA-N EDTA Chemical compound OC(=O)CN(CC(O)=O)CCN(CC(O)=O)CC(O)=O KCXVZYZYPLLWCC-UHFFFAOYSA-N 0.000 description 3
- 101150002621 EPO gene Proteins 0.000 description 3
- PEDCQBHIVMGVHV-UHFFFAOYSA-N Glycerine Chemical compound OCC(O)CO PEDCQBHIVMGVHV-UHFFFAOYSA-N 0.000 description 3
- AJHCSUXXECOXOY-UHFFFAOYSA-N N-glycyl-L-tryptophan Natural products C1=CC=C2C(CC(NC(=O)CN)C(O)=O)=CNC2=C1 AJHCSUXXECOXOY-UHFFFAOYSA-N 0.000 description 3
- 239000007983 Tris buffer Substances 0.000 description 3
- 108010047495 alanylglycine Proteins 0.000 description 3
- 210000000170 cell membrane Anatomy 0.000 description 3
- 238000010367 cloning Methods 0.000 description 3
- 238000010276 construction Methods 0.000 description 3
- 238000001784 detoxification Methods 0.000 description 3
- 238000010790 dilution Methods 0.000 description 3
- 239000012895 dilution Substances 0.000 description 3
- 229940079593 drug Drugs 0.000 description 3
- 239000003814 drug Substances 0.000 description 3
- 238000000605 extraction Methods 0.000 description 3
- 230000028996 humoral immune response Effects 0.000 description 3
- 230000005847 immunogenicity Effects 0.000 description 3
- 208000015181 infectious disease Diseases 0.000 description 3
- 108020004999 messenger RNA Proteins 0.000 description 3
- 108010056582 methionylglutamic acid Proteins 0.000 description 3
- 108010012581 phenylalanylglutamate Proteins 0.000 description 3
- 244000144977 poultry Species 0.000 description 3
- 235000013594 poultry meat Nutrition 0.000 description 3
- 230000008569 process Effects 0.000 description 3
- 108091008146 restriction endonucleases Proteins 0.000 description 3
- 238000004448 titration Methods 0.000 description 3
- LENZDBCJOHFCAS-UHFFFAOYSA-N tris Chemical compound OCC(N)(CO)CO LENZDBCJOHFCAS-UHFFFAOYSA-N 0.000 description 3
- FWMNVWWHGCHHJJ-SKKKGAJSSA-N 4-amino-1-[(2r)-6-amino-2-[[(2r)-2-[[(2r)-2-[[(2r)-2-amino-3-phenylpropanoyl]amino]-3-phenylpropanoyl]amino]-4-methylpentanoyl]amino]hexanoyl]piperidine-4-carboxylic acid Chemical compound C([C@H](C(=O)N[C@H](CC(C)C)C(=O)N[C@H](CCCCN)C(=O)N1CCC(N)(CC1)C(O)=O)NC(=O)[C@H](N)CC=1C=CC=CC=1)C1=CC=CC=C1 FWMNVWWHGCHHJJ-SKKKGAJSSA-N 0.000 description 2
- 208000030507 AIDS Diseases 0.000 description 2
- MFMDKJIPHSWSBM-GUBZILKMSA-N Ala-Lys-Glu Chemical compound [H]N[C@@H](C)C(=O)N[C@@H](CCCCN)C(=O)N[C@@H](CCC(O)=O)C(O)=O MFMDKJIPHSWSBM-GUBZILKMSA-N 0.000 description 2
- NCQMBSJGJMYKCK-ZLUOBGJFSA-N Ala-Ser-Ser Chemical compound [H]N[C@@H](C)C(=O)N[C@@H](CO)C(=O)N[C@@H](CO)C(O)=O NCQMBSJGJMYKCK-ZLUOBGJFSA-N 0.000 description 2
- PQWTZSNVWSOFFK-FXQIFTODSA-N Arg-Asp-Asn Chemical compound C(C[C@@H](C(=O)N[C@@H](CC(=O)O)C(=O)N[C@@H](CC(=O)N)C(=O)O)N)CN=C(N)N PQWTZSNVWSOFFK-FXQIFTODSA-N 0.000 description 2
- RCAUJZASOAFTAJ-FXQIFTODSA-N Arg-Asp-Cys Chemical compound C(C[C@@H](C(=O)N[C@@H](CC(=O)O)C(=O)N[C@@H](CS)C(=O)O)N)CN=C(N)N RCAUJZASOAFTAJ-FXQIFTODSA-N 0.000 description 2
- GMFAGHNRXPSSJS-SRVKXCTJSA-N Arg-Leu-Gln Chemical compound [H]N[C@@H](CCCNC(N)=N)C(=O)N[C@@H](CC(C)C)C(=O)N[C@@H](CCC(N)=O)C(O)=O GMFAGHNRXPSSJS-SRVKXCTJSA-N 0.000 description 2
- KXEGPPNPXOKKHK-ZLUOBGJFSA-N Asn-Asp-Ala Chemical compound [H]N[C@@H](CC(N)=O)C(=O)N[C@@H](CC(O)=O)C(=O)N[C@@H](C)C(O)=O KXEGPPNPXOKKHK-ZLUOBGJFSA-N 0.000 description 2
- 108091003079 Bovine Serum Albumin Proteins 0.000 description 2
- 208000035473 Communicable disease Diseases 0.000 description 2
- 101710121417 Envelope glycoprotein Proteins 0.000 description 2
- WOMUDRVDJMHTCV-DCAQKATOSA-N Glu-Arg-Arg Chemical compound [H]N[C@@H](CCC(O)=O)C(=O)N[C@@H](CCCNC(N)=N)C(=O)N[C@@H](CCCNC(N)=N)C(O)=O WOMUDRVDJMHTCV-DCAQKATOSA-N 0.000 description 2
- 239000004471 Glycine Substances 0.000 description 2
- 229920002527 Glycogen Polymers 0.000 description 2
- 108010001336 Horseradish Peroxidase Proteins 0.000 description 2
- 241000713772 Human immunodeficiency virus 1 Species 0.000 description 2
- 241000712431 Influenza A virus Species 0.000 description 2
- 102100034349 Integrase Human genes 0.000 description 2
- CKLJMWTZIZZHCS-REOHCLBHSA-N L-aspartic acid Chemical compound OC(=O)[C@@H](N)CC(O)=O CKLJMWTZIZZHCS-REOHCLBHSA-N 0.000 description 2
- AGPKZVBTJJNPAG-WHFBIAKZSA-N L-isoleucine Chemical compound CC[C@H](C)[C@H](N)C(O)=O AGPKZVBTJJNPAG-WHFBIAKZSA-N 0.000 description 2
- ROHFNLRQFUQHCH-YFKPBYRVSA-N L-leucine Chemical compound CC(C)C[C@H](N)C(O)=O ROHFNLRQFUQHCH-YFKPBYRVSA-N 0.000 description 2
- COLNVLDHVKWLRT-QMMMGPOBSA-N L-phenylalanine Chemical compound OC(=O)[C@@H](N)CC1=CC=CC=C1 COLNVLDHVKWLRT-QMMMGPOBSA-N 0.000 description 2
- OUYCCCASQSFEME-QMMMGPOBSA-N L-tyrosine Chemical compound OC(=O)[C@@H](N)CC1=CC=C(O)C=C1 OUYCCCASQSFEME-QMMMGPOBSA-N 0.000 description 2
- 241000880493 Leptailurus serval Species 0.000 description 2
- FMEICTQWUKNAGC-YUMQZZPRSA-N Leu-Gly-Asn Chemical compound [H]N[C@@H](CC(C)C)C(=O)NCC(=O)N[C@@H](CC(N)=O)C(O)=O FMEICTQWUKNAGC-YUMQZZPRSA-N 0.000 description 2
- MDDUIRLQCYVRDO-NHCYSSNCSA-N Lys-Val-Asn Chemical compound NC(=O)C[C@@H](C(O)=O)NC(=O)[C@H](C(C)C)NC(=O)[C@@H](N)CCCCN MDDUIRLQCYVRDO-NHCYSSNCSA-N 0.000 description 2
- 241000699670 Mus sp. Species 0.000 description 2
- XMBSYZWANAQXEV-UHFFFAOYSA-N N-alpha-L-glutamyl-L-phenylalanine Natural products OC(=O)CCC(N)C(=O)NC(C(O)=O)CC1=CC=CC=C1 XMBSYZWANAQXEV-UHFFFAOYSA-N 0.000 description 2
- 108700026244 Open Reading Frames Proteins 0.000 description 2
- 239000005662 Paraffin oil Substances 0.000 description 2
- PXIPVTKHYLBLMZ-UHFFFAOYSA-N Sodium azide Chemical compound [Na+].[N-]=[N+]=[N-] PXIPVTKHYLBLMZ-UHFFFAOYSA-N 0.000 description 2
- RFKJNTRMXGCKFE-FHWLQOOXSA-N Val-Leu-Trp Chemical compound C1=CC=C2C(C[C@H](NC(=O)[C@@H](NC(=O)[C@@H](N)C(C)C)CC(C)C)C(O)=O)=CNC2=C1 RFKJNTRMXGCKFE-FHWLQOOXSA-N 0.000 description 2
- XXWBHOWRARMUOC-NHCYSSNCSA-N Val-Lys-Asn Chemical compound CC(C)[C@@H](C(=O)N[C@@H](CCCCN)C(=O)N[C@@H](CC(=O)N)C(=O)O)N XXWBHOWRARMUOC-NHCYSSNCSA-N 0.000 description 2
- RYQUMYBMOJYYDK-NHCYSSNCSA-N Val-Pro-Glu Chemical compound CC(C)[C@@H](C(=O)N1CCC[C@H]1C(=O)N[C@@H](CCC(=O)O)C(=O)O)N RYQUMYBMOJYYDK-NHCYSSNCSA-N 0.000 description 2
- KOSRFJWDECSPRO-UHFFFAOYSA-N alpha-L-glutamyl-L-glutamic acid Natural products OC(=O)CCC(N)C(=O)NC(CCC(O)=O)C(O)=O KOSRFJWDECSPRO-UHFFFAOYSA-N 0.000 description 2
- 108010001271 arginyl-glutamyl-arginine Proteins 0.000 description 2
- 108010052670 arginyl-glutamyl-glutamic acid Proteins 0.000 description 2
- 108010069205 aspartyl-phenylalanine Proteins 0.000 description 2
- 210000004369 blood Anatomy 0.000 description 2
- 239000008280 blood Substances 0.000 description 2
- 230000001413 cellular effect Effects 0.000 description 2
- 239000003153 chemical reaction reagent Substances 0.000 description 2
- 239000003795 chemical substances by application Substances 0.000 description 2
- 238000001816 cooling Methods 0.000 description 2
- 238000005520 cutting process Methods 0.000 description 2
- 108010060199 cysteinylproline Proteins 0.000 description 2
- 238000001035 drying Methods 0.000 description 2
- 239000013613 expression plasmid Substances 0.000 description 2
- 239000012091 fetal bovine serum Substances 0.000 description 2
- 239000012530 fluid Substances 0.000 description 2
- 108010006664 gamma-glutamyl-glycyl-glycine Proteins 0.000 description 2
- 108010057083 glutamyl-aspartyl-leucine Proteins 0.000 description 2
- 108010055341 glutamyl-glutamic acid Proteins 0.000 description 2
- 229940096919 glycogen Drugs 0.000 description 2
- 108010090037 glycyl-alanyl-isoleucine Proteins 0.000 description 2
- 108010066198 glycyl-leucyl-phenylalanine Proteins 0.000 description 2
- 108010048994 glycyl-tyrosyl-alanine Proteins 0.000 description 2
- 108010050848 glycylleucine Proteins 0.000 description 2
- 108010037850 glycylvaline Proteins 0.000 description 2
- 230000035931 haemagglutination Effects 0.000 description 2
- 108010050343 histidyl-alanyl-glutamine Proteins 0.000 description 2
- 108010028295 histidylhistidine Proteins 0.000 description 2
- 108010092114 histidylphenylalanine Proteins 0.000 description 2
- 108010027338 isoleucylcysteine Proteins 0.000 description 2
- 108010090333 leucyl-lysyl-proline Proteins 0.000 description 2
- 210000003928 nasal cavity Anatomy 0.000 description 2
- 108010073025 phenylalanylphenylalanine Proteins 0.000 description 2
- BASFCYQUMIYNBI-UHFFFAOYSA-N platinum Chemical compound [Pt] BASFCYQUMIYNBI-UHFFFAOYSA-N 0.000 description 2
- 230000002265 prevention Effects 0.000 description 2
- 238000011084 recovery Methods 0.000 description 2
- 150000003839 salts Chemical class 0.000 description 2
- 239000011780 sodium chloride Substances 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
- 239000006228 supernatant Substances 0.000 description 2
- 238000013519 translation Methods 0.000 description 2
- 238000001291 vacuum drying Methods 0.000 description 2
- 108010073969 valyllysine Proteins 0.000 description 2
- 230000009385 viral infection Effects 0.000 description 2
- 238000005406 washing Methods 0.000 description 2
- 108091032973 (ribonucleotides)n+m Proteins 0.000 description 1
- JKMHFZQWWAIEOD-UHFFFAOYSA-N 2-[4-(2-hydroxyethyl)piperazin-1-yl]ethanesulfonic acid Chemical compound OCC[NH+]1CCN(CCS([O-])(=O)=O)CC1 JKMHFZQWWAIEOD-UHFFFAOYSA-N 0.000 description 1
- LBJYAILUMSUTAM-ZLUOBGJFSA-N Ala-Asn-Asn Chemical compound [H]N[C@@H](C)C(=O)N[C@@H](CC(N)=O)C(=O)N[C@@H](CC(N)=O)C(O)=O LBJYAILUMSUTAM-ZLUOBGJFSA-N 0.000 description 1
- LSLIRHLIUDVNBN-CIUDSAMLSA-N Ala-Asp-Lys Chemical compound C[C@H](N)C(=O)N[C@@H](CC(O)=O)C(=O)N[C@H](C(O)=O)CCCCN LSLIRHLIUDVNBN-CIUDSAMLSA-N 0.000 description 1
- NKJBKNVQHBZUIX-ACZMJKKPSA-N Ala-Gln-Asp Chemical compound C[C@H](N)C(=O)N[C@@H](CCC(N)=O)C(=O)N[C@@H](CC(O)=O)C(O)=O NKJBKNVQHBZUIX-ACZMJKKPSA-N 0.000 description 1
- BGNLUHXLSAQYRQ-FXQIFTODSA-N Ala-Glu-Gln Chemical compound C[C@H](N)C(=O)N[C@@H](CCC(O)=O)C(=O)N[C@@H](CCC(N)=O)C(O)=O BGNLUHXLSAQYRQ-FXQIFTODSA-N 0.000 description 1
- QHASENCZLDHBGX-ONGXEEELSA-N Ala-Gly-Phe Chemical compound C[C@H](N)C(=O)NCC(=O)N[C@H](C(O)=O)CC1=CC=CC=C1 QHASENCZLDHBGX-ONGXEEELSA-N 0.000 description 1
- PNALXAODQKTNLV-JBDRJPRFSA-N Ala-Ile-Ala Chemical compound C[C@H](N)C(=O)N[C@@H]([C@@H](C)CC)C(=O)N[C@@H](C)C(O)=O PNALXAODQKTNLV-JBDRJPRFSA-N 0.000 description 1
- IFKQPMZRDQZSHI-GHCJXIJMSA-N Ala-Ile-Asn Chemical compound [H]N[C@@H](C)C(=O)N[C@@H]([C@@H](C)CC)C(=O)N[C@@H](CC(N)=O)C(O)=O IFKQPMZRDQZSHI-GHCJXIJMSA-N 0.000 description 1
- XCZXVTHYGSMQGH-NAKRPEOUSA-N Ala-Ile-Met Chemical compound C[C@H]([NH3+])C(=O)N[C@@H]([C@@H](C)CC)C(=O)N[C@@H](CCSC)C([O-])=O XCZXVTHYGSMQGH-NAKRPEOUSA-N 0.000 description 1
- LNNSWWRRYJLGNI-NAKRPEOUSA-N Ala-Ile-Val Chemical compound C[C@H](N)C(=O)N[C@@H]([C@@H](C)CC)C(=O)N[C@@H](C(C)C)C(O)=O LNNSWWRRYJLGNI-NAKRPEOUSA-N 0.000 description 1
- IORKCNUBHNIMKY-CIUDSAMLSA-N Ala-Pro-Glu Chemical compound C[C@H](N)C(=O)N1CCC[C@H]1C(=O)N[C@@H](CCC(O)=O)C(O)=O IORKCNUBHNIMKY-CIUDSAMLSA-N 0.000 description 1
- XAXMJQUMRJAFCH-CQDKDKBSSA-N Ala-Tyr-Lys Chemical compound NCCCC[C@@H](C(O)=O)NC(=O)[C@@H](NC(=O)[C@@H](N)C)CC1=CC=C(O)C=C1 XAXMJQUMRJAFCH-CQDKDKBSSA-N 0.000 description 1
- ZXKNLCPUNZPFGY-LEWSCRJBSA-N Ala-Tyr-Pro Chemical compound C[C@@H](C(=O)N[C@@H](CC1=CC=C(C=C1)O)C(=O)N2CCC[C@@H]2C(=O)O)N ZXKNLCPUNZPFGY-LEWSCRJBSA-N 0.000 description 1
- XPSGESXVBSQZPL-SRVKXCTJSA-N Arg-Arg-Arg Chemical compound NC(N)=NCCC[C@H](N)C(=O)N[C@@H](CCCN=C(N)N)C(=O)N[C@@H](CCCN=C(N)N)C(O)=O XPSGESXVBSQZPL-SRVKXCTJSA-N 0.000 description 1
- ITVINTQUZMQWJR-QXEWZRGKSA-N Arg-Asn-Val Chemical compound [H]N[C@@H](CCCNC(N)=N)C(=O)N[C@@H](CC(N)=O)C(=O)N[C@@H](C(C)C)C(O)=O ITVINTQUZMQWJR-QXEWZRGKSA-N 0.000 description 1
- PNQWAUXQDBIJDY-GUBZILKMSA-N Arg-Glu-Glu Chemical compound [H]N[C@@H](CCCNC(N)=N)C(=O)N[C@@H](CCC(O)=O)C(=O)N[C@@H](CCC(O)=O)C(O)=O PNQWAUXQDBIJDY-GUBZILKMSA-N 0.000 description 1
- OQCWXQJLCDPRHV-UWVGGRQHSA-N Arg-Gly-Leu Chemical compound [H]N[C@@H](CCCNC(N)=N)C(=O)NCC(=O)N[C@@H](CC(C)C)C(O)=O OQCWXQJLCDPRHV-UWVGGRQHSA-N 0.000 description 1
- HCIUUZGFTDTEGM-NAKRPEOUSA-N Arg-Ile-Cys Chemical compound CC[C@H](C)[C@@H](C(=O)N[C@@H](CS)C(=O)O)NC(=O)[C@H](CCCN=C(N)N)N HCIUUZGFTDTEGM-NAKRPEOUSA-N 0.000 description 1
- AGVNTAUPLWIQEN-ZPFDUUQYSA-N Arg-Ile-Glu Chemical compound CC[C@H](C)[C@@H](C(=O)N[C@@H](CCC(=O)O)C(=O)O)NC(=O)[C@H](CCCN=C(N)N)N AGVNTAUPLWIQEN-ZPFDUUQYSA-N 0.000 description 1
- UHFUZWSZQKMDSX-DCAQKATOSA-N Arg-Leu-Asn Chemical compound CC(C)C[C@@H](C(=O)N[C@@H](CC(=O)N)C(=O)O)NC(=O)[C@H](CCCN=C(N)N)N UHFUZWSZQKMDSX-DCAQKATOSA-N 0.000 description 1
- BTJVOUQWFXABOI-IHRRRGAJSA-N Arg-Lys-Lys Chemical compound NCCCC[C@@H](C(O)=O)NC(=O)[C@H](CCCCN)NC(=O)[C@@H](N)CCCNC(N)=N BTJVOUQWFXABOI-IHRRRGAJSA-N 0.000 description 1
- VIINVRPKMUZYOI-DCAQKATOSA-N Arg-Met-Glu Chemical compound [H]N[C@@H](CCCNC(N)=N)C(=O)N[C@@H](CCSC)C(=O)N[C@@H](CCC(O)=O)C(O)=O VIINVRPKMUZYOI-DCAQKATOSA-N 0.000 description 1
- FRBAHXABMQXSJQ-FXQIFTODSA-N Arg-Ser-Ser Chemical compound [H]N[C@@H](CCCNC(N)=N)C(=O)N[C@@H](CO)C(=O)N[C@@H](CO)C(O)=O FRBAHXABMQXSJQ-FXQIFTODSA-N 0.000 description 1
- BECXEHHOZNFFFX-IHRRRGAJSA-N Arg-Ser-Tyr Chemical compound [H]N[C@@H](CCCNC(N)=N)C(=O)N[C@@H](CO)C(=O)N[C@@H](CC1=CC=C(O)C=C1)C(O)=O BECXEHHOZNFFFX-IHRRRGAJSA-N 0.000 description 1
- RYQSYXFGFOTJDJ-RHYQMDGZSA-N Arg-Thr-Leu Chemical compound [H]N[C@@H](CCCNC(N)=N)C(=O)N[C@@H]([C@@H](C)O)C(=O)N[C@@H](CC(C)C)C(O)=O RYQSYXFGFOTJDJ-RHYQMDGZSA-N 0.000 description 1
- 239000004475 Arginine Substances 0.000 description 1
- LEFKSBYHUGUWLP-ACZMJKKPSA-N Asn-Ala-Glu Chemical compound [H]N[C@@H](CC(N)=O)C(=O)N[C@@H](C)C(=O)N[C@@H](CCC(O)=O)C(O)=O LEFKSBYHUGUWLP-ACZMJKKPSA-N 0.000 description 1
- MFFOYNGMOYFPBD-DCAQKATOSA-N Asn-Arg-Leu Chemical compound [H]N[C@@H](CC(N)=O)C(=O)N[C@@H](CCCNC(N)=N)C(=O)N[C@@H](CC(C)C)C(O)=O MFFOYNGMOYFPBD-DCAQKATOSA-N 0.000 description 1
- DAPLJWATMAXPPZ-CIUDSAMLSA-N Asn-Asn-Leu Chemical compound CC(C)C[C@@H](C(O)=O)NC(=O)[C@H](CC(N)=O)NC(=O)[C@@H](N)CC(N)=O DAPLJWATMAXPPZ-CIUDSAMLSA-N 0.000 description 1
- KXFCBAHYSLJCCY-ZLUOBGJFSA-N Asn-Asn-Ser Chemical compound [H]N[C@@H](CC(N)=O)C(=O)N[C@@H](CC(N)=O)C(=O)N[C@@H](CO)C(O)=O KXFCBAHYSLJCCY-ZLUOBGJFSA-N 0.000 description 1
- VKCOHFFSTKCXEQ-OLHMAJIHSA-N Asn-Asn-Thr Chemical compound [H]N[C@@H](CC(N)=O)C(=O)N[C@@H](CC(N)=O)C(=O)N[C@@H]([C@@H](C)O)C(O)=O VKCOHFFSTKCXEQ-OLHMAJIHSA-N 0.000 description 1
- UGXVKHRDGLYFKR-CIUDSAMLSA-N Asn-Asp-Leu Chemical compound CC(C)C[C@@H](C(O)=O)NC(=O)[C@H](CC(O)=O)NC(=O)[C@@H](N)CC(N)=O UGXVKHRDGLYFKR-CIUDSAMLSA-N 0.000 description 1
- HLTLEIXYIJDFOY-ZLUOBGJFSA-N Asn-Cys-Asn Chemical compound NC(=O)C[C@H](N)C(=O)N[C@@H](CS)C(=O)N[C@@H](CC(N)=O)C(O)=O HLTLEIXYIJDFOY-ZLUOBGJFSA-N 0.000 description 1
- PPMTUXJSQDNUDE-CIUDSAMLSA-N Asn-Glu-Arg Chemical compound NC(=O)C[C@H](N)C(=O)N[C@@H](CCC(O)=O)C(=O)N[C@H](C(O)=O)CCCN=C(N)N PPMTUXJSQDNUDE-CIUDSAMLSA-N 0.000 description 1
- MECFLTFREHAZLH-ACZMJKKPSA-N Asn-Glu-Cys Chemical compound C(CC(=O)O)[C@@H](C(=O)N[C@@H](CS)C(=O)O)NC(=O)[C@H](CC(=O)N)N MECFLTFREHAZLH-ACZMJKKPSA-N 0.000 description 1
- DXVMJJNAOVECBA-WHFBIAKZSA-N Asn-Gly-Asn Chemical compound NC(=O)C[C@H](N)C(=O)NCC(=O)N[C@@H](CC(N)=O)C(O)=O DXVMJJNAOVECBA-WHFBIAKZSA-N 0.000 description 1
- PLVAAIPKSGUXDV-WHFBIAKZSA-N Asn-Gly-Cys Chemical compound C([C@@H](C(=O)NCC(=O)N[C@@H](CS)C(=O)O)N)C(=O)N PLVAAIPKSGUXDV-WHFBIAKZSA-N 0.000 description 1
- WONGRTVAMHFGBE-WDSKDSINSA-N Asn-Gly-Gln Chemical compound C(CC(=O)N)[C@@H](C(=O)O)NC(=O)CNC(=O)[C@H](CC(=O)N)N WONGRTVAMHFGBE-WDSKDSINSA-N 0.000 description 1
- FTCGGKNCJZOPNB-WHFBIAKZSA-N Asn-Gly-Ser Chemical compound NC(=O)C[C@H](N)C(=O)NCC(=O)N[C@@H](CO)C(O)=O FTCGGKNCJZOPNB-WHFBIAKZSA-N 0.000 description 1
- RAQMSGVCGSJKCL-FOHZUACHSA-N Asn-Gly-Thr Chemical compound C[C@@H](O)[C@@H](C(O)=O)NC(=O)CNC(=O)[C@@H](N)CC(N)=O RAQMSGVCGSJKCL-FOHZUACHSA-N 0.000 description 1
- OOWSBIOUKIUWLO-RCOVLWMOSA-N Asn-Gly-Val Chemical compound [H]N[C@@H](CC(N)=O)C(=O)NCC(=O)N[C@@H](C(C)C)C(O)=O OOWSBIOUKIUWLO-RCOVLWMOSA-N 0.000 description 1
- XLHLPYFMXGOASD-CIUDSAMLSA-N Asn-His-Asp Chemical compound C1=C(NC=N1)C[C@@H](C(=O)N[C@@H](CC(=O)O)C(=O)O)NC(=O)[C@H](CC(=O)N)N XLHLPYFMXGOASD-CIUDSAMLSA-N 0.000 description 1
- JGIAYNNXZKKKOW-KKUMJFAQSA-N Asn-His-Phe Chemical compound C1=CC=C(C=C1)C[C@@H](C(=O)O)NC(=O)[C@H](CC2=CN=CN2)NC(=O)[C@H](CC(=O)N)N JGIAYNNXZKKKOW-KKUMJFAQSA-N 0.000 description 1
- KMCRKVOLRCOMBG-DJFWLOJKSA-N Asn-Ile-His Chemical compound CC[C@H](C)[C@@H](C(=O)N[C@@H](CC1=CN=CN1)C(=O)O)NC(=O)[C@H](CC(=O)N)N KMCRKVOLRCOMBG-DJFWLOJKSA-N 0.000 description 1
- ZMUQQMGITUJQTI-CIUDSAMLSA-N Asn-Leu-Asn Chemical compound [H]N[C@@H](CC(N)=O)C(=O)N[C@@H](CC(C)C)C(=O)N[C@@H](CC(N)=O)C(O)=O ZMUQQMGITUJQTI-CIUDSAMLSA-N 0.000 description 1
- HDHZCEDPLTVHFZ-GUBZILKMSA-N Asn-Leu-Glu Chemical compound [H]N[C@@H](CC(N)=O)C(=O)N[C@@H](CC(C)C)C(=O)N[C@@H](CCC(O)=O)C(O)=O HDHZCEDPLTVHFZ-GUBZILKMSA-N 0.000 description 1
- TZFQICWZWFNIKU-KKUMJFAQSA-N Asn-Leu-Tyr Chemical compound NC(=O)C[C@H](N)C(=O)N[C@@H](CC(C)C)C(=O)N[C@H](C(O)=O)CC1=CC=C(O)C=C1 TZFQICWZWFNIKU-KKUMJFAQSA-N 0.000 description 1
- NLDNNZKUSLAYFW-NHCYSSNCSA-N Asn-Lys-Val Chemical compound [H]N[C@@H](CC(N)=O)C(=O)N[C@@H](CCCCN)C(=O)N[C@@H](C(C)C)C(O)=O NLDNNZKUSLAYFW-NHCYSSNCSA-N 0.000 description 1
- IDUUACUJKUXKKD-VEVYYDQMSA-N Asn-Pro-Thr Chemical compound [H]N[C@@H](CC(N)=O)C(=O)N1CCC[C@H]1C(=O)N[C@@H]([C@@H](C)O)C(O)=O IDUUACUJKUXKKD-VEVYYDQMSA-N 0.000 description 1
- XTMZYFMTYJNABC-ZLUOBGJFSA-N Asn-Ser-Ala Chemical compound C[C@@H](C(=O)O)NC(=O)[C@H](CO)NC(=O)[C@H](CC(=O)N)N XTMZYFMTYJNABC-ZLUOBGJFSA-N 0.000 description 1
- NPZJLGMWMDNQDD-GHCJXIJMSA-N Asn-Ser-Ile Chemical compound [H]N[C@@H](CC(N)=O)C(=O)N[C@@H](CO)C(=O)N[C@@H]([C@@H](C)CC)C(O)=O NPZJLGMWMDNQDD-GHCJXIJMSA-N 0.000 description 1
- WUQXMTITJLFXAU-JIOCBJNQSA-N Asn-Thr-Pro Chemical compound C[C@H]([C@@H](C(=O)N1CCC[C@@H]1C(=O)O)NC(=O)[C@H](CC(=O)N)N)O WUQXMTITJLFXAU-JIOCBJNQSA-N 0.000 description 1
- LMIWYCWRJVMAIQ-NHCYSSNCSA-N Asn-Val-Lys Chemical compound CC(C)[C@@H](C(=O)N[C@@H](CCCCN)C(=O)O)NC(=O)[C@H](CC(=O)N)N LMIWYCWRJVMAIQ-NHCYSSNCSA-N 0.000 description 1
- XOQYDFCQPWAMSA-KKHAAJSZSA-N Asn-Val-Thr Chemical compound [H]N[C@@H](CC(N)=O)C(=O)N[C@@H](C(C)C)C(=O)N[C@@H]([C@@H](C)O)C(O)=O XOQYDFCQPWAMSA-KKHAAJSZSA-N 0.000 description 1
- WQAOZCVOOYUWKG-LSJOCFKGSA-N Asn-Val-Val Chemical compound CC(C)[C@@H](C(=O)N[C@@H](C(C)C)C(=O)O)NC(=O)[C@H](CC(=O)N)N WQAOZCVOOYUWKG-LSJOCFKGSA-N 0.000 description 1
- KRXIWXCXOARFNT-ZLUOBGJFSA-N Asp-Ala-Ala Chemical compound OC(=O)[C@H](C)NC(=O)[C@H](C)NC(=O)[C@@H](N)CC(O)=O KRXIWXCXOARFNT-ZLUOBGJFSA-N 0.000 description 1
- XBQSLMACWDXWLJ-GHCJXIJMSA-N Asp-Ala-Ile Chemical compound [H]N[C@@H](CC(O)=O)C(=O)N[C@@H](C)C(=O)N[C@@H]([C@@H](C)CC)C(O)=O XBQSLMACWDXWLJ-GHCJXIJMSA-N 0.000 description 1
- PMEHKVHZQKJACS-PEFMBERDSA-N Asp-Gln-Ile Chemical compound [H]N[C@@H](CC(O)=O)C(=O)N[C@@H](CCC(N)=O)C(=O)N[C@@H]([C@@H](C)CC)C(O)=O PMEHKVHZQKJACS-PEFMBERDSA-N 0.000 description 1
- OVPHVTCDVYYTHN-AVGNSLFASA-N Asp-Glu-Phe Chemical compound OC(=O)C[C@H](N)C(=O)N[C@@H](CCC(O)=O)C(=O)N[C@H](C(O)=O)CC1=CC=CC=C1 OVPHVTCDVYYTHN-AVGNSLFASA-N 0.000 description 1
- NRIFEOUAFLTMFJ-AAEUAGOBSA-N Asp-Gly-Trp Chemical compound [H]N[C@@H](CC(O)=O)C(=O)NCC(=O)N[C@@H](CC1=CNC2=C1C=CC=C2)C(O)=O NRIFEOUAFLTMFJ-AAEUAGOBSA-N 0.000 description 1
- HOBNTSHITVVNBN-ZPFDUUQYSA-N Asp-Ile-Leu Chemical compound CC[C@H](C)[C@@H](C(=O)N[C@@H](CC(C)C)C(=O)O)NC(=O)[C@H](CC(=O)O)N HOBNTSHITVVNBN-ZPFDUUQYSA-N 0.000 description 1
- VSMYBNPOHYAXSD-GUBZILKMSA-N Asp-Lys-Glu Chemical compound OC(=O)C[C@H](N)C(=O)N[C@@H](CCCCN)C(=O)N[C@@H](CCC(O)=O)C(O)=O VSMYBNPOHYAXSD-GUBZILKMSA-N 0.000 description 1
- MYLZFUMPZCPJCJ-NHCYSSNCSA-N Asp-Lys-Val Chemical compound [H]N[C@@H](CC(O)=O)C(=O)N[C@@H](CCCCN)C(=O)N[C@@H](C(C)C)C(O)=O MYLZFUMPZCPJCJ-NHCYSSNCSA-N 0.000 description 1
- KRQFMDNIUOVRIF-KKUMJFAQSA-N Asp-Phe-His Chemical compound C1=CC=C(C=C1)C[C@@H](C(=O)N[C@@H](CC2=CN=CN2)C(=O)O)NC(=O)[C@H](CC(=O)O)N KRQFMDNIUOVRIF-KKUMJFAQSA-N 0.000 description 1
- WMLFFCRUSPNENW-ZLUOBGJFSA-N Asp-Ser-Ala Chemical compound [H]N[C@@H](CC(O)=O)C(=O)N[C@@H](CO)C(=O)N[C@@H](C)C(O)=O WMLFFCRUSPNENW-ZLUOBGJFSA-N 0.000 description 1
- XXAMCEGRCZQGEM-ZLUOBGJFSA-N Asp-Ser-Asn Chemical compound [H]N[C@@H](CC(O)=O)C(=O)N[C@@H](CO)C(=O)N[C@@H](CC(N)=O)C(O)=O XXAMCEGRCZQGEM-ZLUOBGJFSA-N 0.000 description 1
- KBJVTFWQWXCYCQ-IUKAMOBKSA-N Asp-Thr-Ile Chemical compound [H]N[C@@H](CC(O)=O)C(=O)N[C@@H]([C@@H](C)O)C(=O)N[C@@H]([C@@H](C)CC)C(O)=O KBJVTFWQWXCYCQ-IUKAMOBKSA-N 0.000 description 1
- KNDCWFXCFKSEBM-AVGNSLFASA-N Asp-Tyr-Glu Chemical compound [H]N[C@@H](CC(O)=O)C(=O)N[C@@H](CC1=CC=C(O)C=C1)C(=O)N[C@@H](CCC(O)=O)C(O)=O KNDCWFXCFKSEBM-AVGNSLFASA-N 0.000 description 1
- 241000271566 Aves Species 0.000 description 1
- 241000894006 Bacteria Species 0.000 description 1
- COXVTLYNGOIATD-HVMBLDELSA-N CC1=C(C=CC(=C1)C1=CC(C)=C(C=C1)\N=N\C1=C(O)C2=C(N)C(=CC(=C2C=C1)S(O)(=O)=O)S(O)(=O)=O)\N=N\C1=CC=C2C(=CC(=C(N)C2=C1O)S(O)(=O)=O)S(O)(=O)=O Chemical compound CC1=C(C=CC(=C1)C1=CC(C)=C(C=C1)\N=N\C1=C(O)C2=C(N)C(=CC(=C2C=C1)S(O)(=O)=O)S(O)(=O)=O)\N=N\C1=CC=C2C(=CC(=C(N)C2=C1O)S(O)(=O)=O)S(O)(=O)=O COXVTLYNGOIATD-HVMBLDELSA-N 0.000 description 1
- 101100008050 Caenorhabditis elegans cut-6 gene Proteins 0.000 description 1
- 241000282472 Canis lupus familiaris Species 0.000 description 1
- 241000282693 Cercopithecidae Species 0.000 description 1
- 101100358659 Chlorobium chlorochromatii (strain CaD3) rplY gene Proteins 0.000 description 1
- 241000711573 Coronaviridae Species 0.000 description 1
- LHLSSZYQFUNWRZ-NAKRPEOUSA-N Cys-Arg-Ile Chemical compound [H]N[C@@H](CS)C(=O)N[C@@H](CCCNC(N)=N)C(=O)N[C@@H]([C@@H](C)CC)C(O)=O LHLSSZYQFUNWRZ-NAKRPEOUSA-N 0.000 description 1
- NQSUTVRXXBGVDQ-LKXGYXEUSA-N Cys-Asn-Thr Chemical compound [H]N[C@@H](CS)C(=O)N[C@@H](CC(N)=O)C(=O)N[C@@H]([C@@H](C)O)C(O)=O NQSUTVRXXBGVDQ-LKXGYXEUSA-N 0.000 description 1
- UWXFFVQPAMBETM-ZLUOBGJFSA-N Cys-Asp-Asn Chemical compound [H]N[C@@H](CS)C(=O)N[C@@H](CC(O)=O)C(=O)N[C@@H](CC(N)=O)C(O)=O UWXFFVQPAMBETM-ZLUOBGJFSA-N 0.000 description 1
- XABFFGOGKOORCG-CIUDSAMLSA-N Cys-Asp-Leu Chemical compound [H]N[C@@H](CS)C(=O)N[C@@H](CC(O)=O)C(=O)N[C@@H](CC(C)C)C(O)=O XABFFGOGKOORCG-CIUDSAMLSA-N 0.000 description 1
- ZMWOJVAXTOUHAP-ZKWXMUAHSA-N Cys-Ile-Gly Chemical compound CC[C@H](C)[C@@H](C(=O)NCC(=O)O)NC(=O)[C@H](CS)N ZMWOJVAXTOUHAP-ZKWXMUAHSA-N 0.000 description 1
- JUNZLDGUJZIUCO-IHRRRGAJSA-N Cys-Pro-Tyr Chemical compound C1C[C@H](N(C1)C(=O)[C@H](CS)N)C(=O)N[C@@H](CC2=CC=C(C=C2)O)C(=O)O JUNZLDGUJZIUCO-IHRRRGAJSA-N 0.000 description 1
- KVCJEMHFLGVINV-ZLUOBGJFSA-N Cys-Ser-Asn Chemical compound SC[C@H](N)C(=O)N[C@@H](CO)C(=O)N[C@H](C(O)=O)CC(N)=O KVCJEMHFLGVINV-ZLUOBGJFSA-N 0.000 description 1
- CLEFUAZULXANBU-MELADBBJSA-N Cys-Tyr-Pro Chemical compound C1C[C@@H](N(C1)C(=O)[C@H](CC2=CC=C(C=C2)O)NC(=O)[C@H](CS)N)C(=O)O CLEFUAZULXANBU-MELADBBJSA-N 0.000 description 1
- 102000012410 DNA Ligases Human genes 0.000 description 1
- 108010061982 DNA Ligases Proteins 0.000 description 1
- 230000004544 DNA amplification Effects 0.000 description 1
- 108010014303 DNA-directed DNA polymerase Proteins 0.000 description 1
- 102000016928 DNA-directed DNA polymerase Human genes 0.000 description 1
- 101150029662 E1 gene Proteins 0.000 description 1
- 101150013359 E7 gene Proteins 0.000 description 1
- 108090000394 Erythropoietin Proteins 0.000 description 1
- 241000588724 Escherichia coli Species 0.000 description 1
- LZRMPXRYLLTAJX-GUBZILKMSA-N Gln-Arg-Glu Chemical compound [H]N[C@@H](CCC(N)=O)C(=O)N[C@@H](CCCNC(N)=N)C(=O)N[C@@H](CCC(O)=O)C(O)=O LZRMPXRYLLTAJX-GUBZILKMSA-N 0.000 description 1
- PRBLYKYHAJEABA-SRVKXCTJSA-N Gln-Arg-Leu Chemical compound [H]N[C@@H](CCC(N)=O)C(=O)N[C@@H](CCCNC(N)=N)C(=O)N[C@@H](CC(C)C)C(O)=O PRBLYKYHAJEABA-SRVKXCTJSA-N 0.000 description 1
- CGVWDTRDPLOMHZ-FXQIFTODSA-N Gln-Glu-Asp Chemical compound [H]N[C@@H](CCC(N)=O)C(=O)N[C@@H](CCC(O)=O)C(=O)N[C@@H](CC(O)=O)C(O)=O CGVWDTRDPLOMHZ-FXQIFTODSA-N 0.000 description 1
- QQAPDATZKKTBIY-YUMQZZPRSA-N Gln-Gly-Met Chemical compound [H]N[C@@H](CCC(N)=O)C(=O)NCC(=O)N[C@@H](CCSC)C(O)=O QQAPDATZKKTBIY-YUMQZZPRSA-N 0.000 description 1
- DAAUVRPSZRDMBV-KBIXCLLPSA-N Gln-Ile-Cys Chemical compound CC[C@H](C)[C@@H](C(=O)N[C@@H](CS)C(=O)O)NC(=O)[C@H](CCC(=O)N)N DAAUVRPSZRDMBV-KBIXCLLPSA-N 0.000 description 1
- FTIJVMLAGRAYMJ-MNXVOIDGSA-N Gln-Ile-Leu Chemical compound CC(C)C[C@@H](C(O)=O)NC(=O)[C@H]([C@@H](C)CC)NC(=O)[C@@H](N)CCC(N)=O FTIJVMLAGRAYMJ-MNXVOIDGSA-N 0.000 description 1
- IHSGESFHTMFHRB-GUBZILKMSA-N Gln-Lys-Ala Chemical compound OC(=O)[C@H](C)NC(=O)[C@H](CCCCN)NC(=O)[C@@H](N)CCC(N)=O IHSGESFHTMFHRB-GUBZILKMSA-N 0.000 description 1
- QMVCEWKHIUHTSD-GUBZILKMSA-N Gln-Met-Glu Chemical compound CSCC[C@@H](C(=O)N[C@@H](CCC(=O)O)C(=O)O)NC(=O)[C@H](CCC(=O)N)N QMVCEWKHIUHTSD-GUBZILKMSA-N 0.000 description 1
- VLOLPWWCNKWRNB-LOKLDPHHSA-N Gln-Thr-Pro Chemical compound C[C@H]([C@@H](C(=O)N1CCC[C@@H]1C(=O)O)NC(=O)[C@H](CCC(=O)N)N)O VLOLPWWCNKWRNB-LOKLDPHHSA-N 0.000 description 1
- VDMABHYXBULDGN-LAEOZQHASA-N Gln-Val-Asp Chemical compound [H]N[C@@H](CCC(N)=O)C(=O)N[C@@H](C(C)C)C(=O)N[C@@H](CC(O)=O)C(O)=O VDMABHYXBULDGN-LAEOZQHASA-N 0.000 description 1
- FHPXTPQBODWBIY-CIUDSAMLSA-N Glu-Ala-Arg Chemical compound [H]N[C@@H](CCC(O)=O)C(=O)N[C@@H](C)C(=O)N[C@@H](CCCNC(N)=N)C(O)=O FHPXTPQBODWBIY-CIUDSAMLSA-N 0.000 description 1
- NCWOMXABNYEPLY-NRPADANISA-N Glu-Ala-Val Chemical compound [H]N[C@@H](CCC(O)=O)C(=O)N[C@@H](C)C(=O)N[C@@H](C(C)C)C(O)=O NCWOMXABNYEPLY-NRPADANISA-N 0.000 description 1
- YYOBUPFZLKQUAX-FXQIFTODSA-N Glu-Asn-Glu Chemical compound [H]N[C@@H](CCC(O)=O)C(=O)N[C@@H](CC(N)=O)C(=O)N[C@@H](CCC(O)=O)C(O)=O YYOBUPFZLKQUAX-FXQIFTODSA-N 0.000 description 1
- DSPQRJXOIXHOHK-WDSKDSINSA-N Glu-Asp-Gly Chemical compound OC(=O)CC[C@H](N)C(=O)N[C@@H](CC(O)=O)C(=O)NCC(O)=O DSPQRJXOIXHOHK-WDSKDSINSA-N 0.000 description 1
- JVSBYEDSSRZQGV-GUBZILKMSA-N Glu-Asp-Leu Chemical compound CC(C)C[C@@H](C(O)=O)NC(=O)[C@H](CC(O)=O)NC(=O)[C@@H](N)CCC(O)=O JVSBYEDSSRZQGV-GUBZILKMSA-N 0.000 description 1
- ZXLZWUQBRYGDNS-CIUDSAMLSA-N Glu-Cys-Met Chemical compound CSCC[C@@H](C(=O)O)NC(=O)[C@H](CS)NC(=O)[C@H](CCC(=O)O)N ZXLZWUQBRYGDNS-CIUDSAMLSA-N 0.000 description 1
- OWVURWCRZZMAOZ-XHNCKOQMSA-N Glu-Cys-Pro Chemical compound C1C[C@@H](N(C1)C(=O)[C@H](CS)NC(=O)[C@H](CCC(=O)O)N)C(=O)O OWVURWCRZZMAOZ-XHNCKOQMSA-N 0.000 description 1
- PXHABOCPJVTGEK-BQBZGAKWSA-N Glu-Gln-Gly Chemical compound [H]N[C@@H](CCC(O)=O)C(=O)N[C@@H](CCC(N)=O)C(=O)NCC(O)=O PXHABOCPJVTGEK-BQBZGAKWSA-N 0.000 description 1
- VFZIDQZAEBORGY-GLLZPBPUSA-N Glu-Gln-Thr Chemical compound [H]N[C@@H](CCC(O)=O)C(=O)N[C@@H](CCC(N)=O)C(=O)N[C@@H]([C@@H](C)O)C(O)=O VFZIDQZAEBORGY-GLLZPBPUSA-N 0.000 description 1
- AUTNXSQEVVHSJK-YVNDNENWSA-N Glu-Glu-Ile Chemical compound CC[C@H](C)[C@@H](C(O)=O)NC(=O)[C@H](CCC(O)=O)NC(=O)[C@@H](N)CCC(O)=O AUTNXSQEVVHSJK-YVNDNENWSA-N 0.000 description 1
- OAGVHWYIBZMWLA-YFKPBYRVSA-N Glu-Gly-Gly Chemical compound OC(=O)CC[C@H](N)C(=O)NCC(=O)NCC(O)=O OAGVHWYIBZMWLA-YFKPBYRVSA-N 0.000 description 1
- ZWABFSSWTSAMQN-KBIXCLLPSA-N Glu-Ile-Ala Chemical compound [H]N[C@@H](CCC(O)=O)C(=O)N[C@@H]([C@@H](C)CC)C(=O)N[C@@H](C)C(O)=O ZWABFSSWTSAMQN-KBIXCLLPSA-N 0.000 description 1
- IRXNJYPKBVERCW-DCAQKATOSA-N Glu-Leu-Glu Chemical compound [H]N[C@@H](CCC(O)=O)C(=O)N[C@@H](CC(C)C)C(=O)N[C@@H](CCC(O)=O)C(O)=O IRXNJYPKBVERCW-DCAQKATOSA-N 0.000 description 1
- IVGJYOOGJLFKQE-AVGNSLFASA-N Glu-Leu-Lys Chemical compound CC(C)C[C@@H](C(=O)N[C@@H](CCCCN)C(=O)O)NC(=O)[C@H](CCC(=O)O)N IVGJYOOGJLFKQE-AVGNSLFASA-N 0.000 description 1
- OQXDUSZKISQQSS-GUBZILKMSA-N Glu-Lys-Ala Chemical compound [H]N[C@@H](CCC(O)=O)C(=O)N[C@@H](CCCCN)C(=O)N[C@@H](C)C(O)=O OQXDUSZKISQQSS-GUBZILKMSA-N 0.000 description 1
- SWRVAQHFBRZVNX-GUBZILKMSA-N Glu-Lys-Asn Chemical compound [H]N[C@@H](CCC(O)=O)C(=O)N[C@@H](CCCCN)C(=O)N[C@@H](CC(N)=O)C(O)=O SWRVAQHFBRZVNX-GUBZILKMSA-N 0.000 description 1
- QDMVXRNLOPTPIE-WDCWCFNPSA-N Glu-Lys-Thr Chemical compound [H]N[C@@H](CCC(O)=O)C(=O)N[C@@H](CCCCN)C(=O)N[C@@H]([C@@H](C)O)C(O)=O QDMVXRNLOPTPIE-WDCWCFNPSA-N 0.000 description 1
- UERORLSAFUHDGU-AVGNSLFASA-N Glu-Phe-Asn Chemical compound C1=CC=C(C=C1)C[C@@H](C(=O)N[C@@H](CC(=O)N)C(=O)O)NC(=O)[C@H](CCC(=O)O)N UERORLSAFUHDGU-AVGNSLFASA-N 0.000 description 1
- JZJGEKDPWVJOLD-QEWYBTABSA-N Glu-Phe-Ile Chemical compound [H]N[C@@H](CCC(O)=O)C(=O)N[C@@H](CC1=CC=CC=C1)C(=O)N[C@@H]([C@@H](C)CC)C(O)=O JZJGEKDPWVJOLD-QEWYBTABSA-N 0.000 description 1
- MIIGESVJEBDJMP-FHWLQOOXSA-N Glu-Phe-Tyr Chemical compound C([C@H](NC(=O)[C@H](CCC(O)=O)N)C(=O)N[C@@H](CC=1C=CC(O)=CC=1)C(O)=O)C1=CC=CC=C1 MIIGESVJEBDJMP-FHWLQOOXSA-N 0.000 description 1
- ALMBZBOCGSVSAI-ACZMJKKPSA-N Glu-Ser-Asn Chemical compound C(CC(=O)O)[C@@H](C(=O)N[C@@H](CO)C(=O)N[C@@H](CC(=O)N)C(=O)O)N ALMBZBOCGSVSAI-ACZMJKKPSA-N 0.000 description 1
- GUOWMVFLAJNPDY-CIUDSAMLSA-N Glu-Ser-Met Chemical compound [H]N[C@@H](CCC(O)=O)C(=O)N[C@@H](CO)C(=O)N[C@@H](CCSC)C(O)=O GUOWMVFLAJNPDY-CIUDSAMLSA-N 0.000 description 1
- JWNZHMSRZXXGTM-XKBZYTNZSA-N Glu-Ser-Thr Chemical compound [H]N[C@@H](CCC(O)=O)C(=O)N[C@@H](CO)C(=O)N[C@@H]([C@@H](C)O)C(O)=O JWNZHMSRZXXGTM-XKBZYTNZSA-N 0.000 description 1
- OGCIHJPYKVSMTE-YUMQZZPRSA-N Gly-Arg-Glu Chemical compound [H]NCC(=O)N[C@@H](CCCNC(N)=N)C(=O)N[C@@H](CCC(O)=O)C(O)=O OGCIHJPYKVSMTE-YUMQZZPRSA-N 0.000 description 1
- JVACNFOPSUPDTK-QWRGUYRKSA-N Gly-Asn-Phe Chemical compound NCC(=O)N[C@@H](CC(N)=O)C(=O)N[C@H](C(O)=O)CC1=CC=CC=C1 JVACNFOPSUPDTK-QWRGUYRKSA-N 0.000 description 1
- XRTDOIOIBMAXCT-NKWVEPMBSA-N Gly-Asn-Pro Chemical compound C1C[C@@H](N(C1)C(=O)[C@H](CC(=O)N)NC(=O)CN)C(=O)O XRTDOIOIBMAXCT-NKWVEPMBSA-N 0.000 description 1
- RPLLQZBOVIVGMX-QWRGUYRKSA-N Gly-Asp-Phe Chemical compound [H]NCC(=O)N[C@@H](CC(O)=O)C(=O)N[C@@H](CC1=CC=CC=C1)C(O)=O RPLLQZBOVIVGMX-QWRGUYRKSA-N 0.000 description 1
- LCNXZQROPKFGQK-WHFBIAKZSA-N Gly-Asp-Ser Chemical compound NCC(=O)N[C@@H](CC(O)=O)C(=O)N[C@@H](CO)C(O)=O LCNXZQROPKFGQK-WHFBIAKZSA-N 0.000 description 1
- SABZDFAAOJATBR-QWRGUYRKSA-N Gly-Cys-Phe Chemical compound [H]NCC(=O)N[C@@H](CS)C(=O)N[C@@H](CC1=CC=CC=C1)C(O)=O SABZDFAAOJATBR-QWRGUYRKSA-N 0.000 description 1
- GNPVTZJUUBPZKW-WDSKDSINSA-N Gly-Gln-Ser Chemical compound [H]NCC(=O)N[C@@H](CCC(N)=O)C(=O)N[C@@H](CO)C(O)=O GNPVTZJUUBPZKW-WDSKDSINSA-N 0.000 description 1
- BIRKKBCSAIHDDF-WDSKDSINSA-N Gly-Glu-Cys Chemical compound NCC(=O)N[C@@H](CCC(O)=O)C(=O)N[C@@H](CS)C(O)=O BIRKKBCSAIHDDF-WDSKDSINSA-N 0.000 description 1
- DENRBIYENOKSEX-PEXQALLHSA-N Gly-Ile-His Chemical compound NCC(=O)N[C@@H]([C@@H](C)CC)C(=O)N[C@H](C(O)=O)CC1=CN=CN1 DENRBIYENOKSEX-PEXQALLHSA-N 0.000 description 1
- NSTUFLGQJCOCDL-UWVGGRQHSA-N Gly-Leu-Arg Chemical compound NCC(=O)N[C@@H](CC(C)C)C(=O)N[C@H](C(O)=O)CCCN=C(N)N NSTUFLGQJCOCDL-UWVGGRQHSA-N 0.000 description 1
- NNCSJUBVFBDDLC-YUMQZZPRSA-N Gly-Leu-Ser Chemical compound NCC(=O)N[C@@H](CC(C)C)C(=O)N[C@@H](CO)C(O)=O NNCSJUBVFBDDLC-YUMQZZPRSA-N 0.000 description 1
- MHXKHKWHPNETGG-QWRGUYRKSA-N Gly-Lys-Leu Chemical compound [H]NCC(=O)N[C@@H](CCCCN)C(=O)N[C@@H](CC(C)C)C(O)=O MHXKHKWHPNETGG-QWRGUYRKSA-N 0.000 description 1
- FJWSJWACLMTDMI-WPRPVWTQSA-N Gly-Met-Val Chemical compound [H]NCC(=O)N[C@@H](CCSC)C(=O)N[C@@H](C(C)C)C(O)=O FJWSJWACLMTDMI-WPRPVWTQSA-N 0.000 description 1
- IBYOLNARKHMLBG-WHOFXGATSA-N Gly-Phe-Ile Chemical compound CC[C@H](C)[C@@H](C(O)=O)NC(=O)[C@@H](NC(=O)CN)CC1=CC=CC=C1 IBYOLNARKHMLBG-WHOFXGATSA-N 0.000 description 1
- SOEGEPHNZOISMT-BYPYZUCNSA-N Gly-Ser-Gly Chemical compound NCC(=O)N[C@@H](CO)C(=O)NCC(O)=O SOEGEPHNZOISMT-BYPYZUCNSA-N 0.000 description 1
- FFALDIDGPLUDKV-ZDLURKLDSA-N Gly-Thr-Ser Chemical compound [H]NCC(=O)N[C@@H]([C@@H](C)O)C(=O)N[C@@H](CO)C(O)=O FFALDIDGPLUDKV-ZDLURKLDSA-N 0.000 description 1
- FOKISINOENBSDM-WLTAIBSBSA-N Gly-Thr-Tyr Chemical compound [H]NCC(=O)N[C@@H]([C@@H](C)O)C(=O)N[C@@H](CC1=CC=C(O)C=C1)C(O)=O FOKISINOENBSDM-WLTAIBSBSA-N 0.000 description 1
- PYFHPYDQHCEVIT-KBPBESRZSA-N Gly-Trp-Gln Chemical compound [H]NCC(=O)N[C@@H](CC1=CNC2=C1C=CC=C2)C(=O)N[C@@H](CCC(N)=O)C(O)=O PYFHPYDQHCEVIT-KBPBESRZSA-N 0.000 description 1
- SFOXOSKVTLDEDM-HOTGVXAUSA-N Gly-Trp-Leu Chemical compound C1=CC=C2C(C[C@@H](C(=O)N[C@@H](CC(C)C)C(O)=O)NC(=O)CN)=CNC2=C1 SFOXOSKVTLDEDM-HOTGVXAUSA-N 0.000 description 1
- KBBFOULZCHWGJX-KBPBESRZSA-N Gly-Tyr-His Chemical compound C1=CC(=CC=C1C[C@@H](C(=O)N[C@@H](CC2=CN=CN2)C(=O)O)NC(=O)CN)O KBBFOULZCHWGJX-KBPBESRZSA-N 0.000 description 1
- FNXSYBOHALPRHV-ONGXEEELSA-N Gly-Val-Lys Chemical compound NCC(=O)N[C@@H](C(C)C)C(=O)N[C@H](C(O)=O)CCCCN FNXSYBOHALPRHV-ONGXEEELSA-N 0.000 description 1
- SBVMXEZQJVUARN-XPUUQOCRSA-N Gly-Val-Ser Chemical compound NCC(=O)N[C@@H](C(C)C)C(=O)N[C@@H](CO)C(O)=O SBVMXEZQJVUARN-XPUUQOCRSA-N 0.000 description 1
- AFMOTCMSEBITOE-YEPSODPASA-N Gly-Val-Thr Chemical compound NCC(=O)N[C@@H](C(C)C)C(=O)N[C@@H]([C@@H](C)O)C(O)=O AFMOTCMSEBITOE-YEPSODPASA-N 0.000 description 1
- JBCLFWXMTIKCCB-UHFFFAOYSA-N H-Gly-Phe-OH Natural products NCC(=O)NC(C(O)=O)CC1=CC=CC=C1 JBCLFWXMTIKCCB-UHFFFAOYSA-N 0.000 description 1
- 239000007995 HEPES buffer Substances 0.000 description 1
- QIVPRLJQQVXCIY-HGNGGELXSA-N His-Ala-Gln Chemical compound C[C@H](NC(=O)[C@@H](N)Cc1cnc[nH]1)C(=O)N[C@@H](CCC(N)=O)C(O)=O QIVPRLJQQVXCIY-HGNGGELXSA-N 0.000 description 1
- NOQPTNXSGNPJNS-YUMQZZPRSA-N His-Asn-Gly Chemical compound [H]N[C@@H](CC1=CNC=N1)C(=O)N[C@@H](CC(N)=O)C(=O)NCC(O)=O NOQPTNXSGNPJNS-YUMQZZPRSA-N 0.000 description 1
- UZZXGLOJRZKYEL-DJFWLOJKSA-N His-Asn-Ile Chemical compound [H]N[C@@H](CC1=CNC=N1)C(=O)N[C@@H](CC(N)=O)C(=O)N[C@@H]([C@@H](C)CC)C(O)=O UZZXGLOJRZKYEL-DJFWLOJKSA-N 0.000 description 1
- VOEGKUNRHYKYSU-XVYDVKMFSA-N His-Asp-Ala Chemical compound [H]N[C@@H](CC1=CNC=N1)C(=O)N[C@@H](CC(O)=O)C(=O)N[C@@H](C)C(O)=O VOEGKUNRHYKYSU-XVYDVKMFSA-N 0.000 description 1
- ZZLWLWSUIBSMNP-CIUDSAMLSA-N His-Asp-Ser Chemical compound [H]N[C@@H](CC1=CNC=N1)C(=O)N[C@@H](CC(O)=O)C(=O)N[C@@H](CO)C(O)=O ZZLWLWSUIBSMNP-CIUDSAMLSA-N 0.000 description 1
- PYNUBZSXKQKAHL-UWVGGRQHSA-N His-Gly-Arg Chemical compound [H]N[C@@H](CC1=CNC=N1)C(=O)NCC(=O)N[C@@H](CCCNC(N)=N)C(O)=O PYNUBZSXKQKAHL-UWVGGRQHSA-N 0.000 description 1
- JBSLJUPMTYLLFH-MELADBBJSA-N His-His-Pro Chemical compound C1C[C@@H](N(C1)C(=O)[C@H](CC2=CN=CN2)NC(=O)[C@H](CC3=CN=CN3)N)C(=O)O JBSLJUPMTYLLFH-MELADBBJSA-N 0.000 description 1
- YAALVYQFVJNXIV-KKUMJFAQSA-N His-Leu-Leu Chemical compound CC(C)C[C@@H](C(O)=O)NC(=O)[C@H](CC(C)C)NC(=O)[C@@H](N)CC1=CN=CN1 YAALVYQFVJNXIV-KKUMJFAQSA-N 0.000 description 1
- GNBHSMFBUNEWCJ-DCAQKATOSA-N His-Pro-Asn Chemical compound [H]N[C@@H](CC1=CNC=N1)C(=O)N1CCC[C@H]1C(=O)N[C@@H](CC(N)=O)C(O)=O GNBHSMFBUNEWCJ-DCAQKATOSA-N 0.000 description 1
- VCBWXASUBZIFLQ-IHRRRGAJSA-N His-Pro-Leu Chemical compound [H]N[C@@H](CC1=CNC=N1)C(=O)N1CCC[C@H]1C(=O)N[C@@H](CC(C)C)C(O)=O VCBWXASUBZIFLQ-IHRRRGAJSA-N 0.000 description 1
- CWSZWFILCNSNEX-CIUDSAMLSA-N His-Ser-Asn Chemical compound C1=C(NC=N1)C[C@@H](C(=O)N[C@@H](CO)C(=O)N[C@@H](CC(=O)N)C(=O)O)N CWSZWFILCNSNEX-CIUDSAMLSA-N 0.000 description 1
- 102000018713 Histocompatibility Antigens Class II Human genes 0.000 description 1
- 108010027412 Histocompatibility Antigens Class II Proteins 0.000 description 1
- 241000282412 Homo Species 0.000 description 1
- 101000987586 Homo sapiens Eosinophil peroxidase Proteins 0.000 description 1
- 101000920686 Homo sapiens Erythropoietin Proteins 0.000 description 1
- 101001094545 Homo sapiens Retrotransposon-like protein 1 Proteins 0.000 description 1
- 241000701806 Human papillomavirus Species 0.000 description 1
- 241000341655 Human papillomavirus type 16 Species 0.000 description 1
- CYHYBSGMHMHKOA-CIQUZCHMSA-N Ile-Ala-Thr Chemical compound CC[C@H](C)[C@@H](C(=O)N[C@@H](C)C(=O)N[C@@H]([C@@H](C)O)C(=O)O)N CYHYBSGMHMHKOA-CIQUZCHMSA-N 0.000 description 1
- ZZHGKECPZXPXJF-PCBIJLKTSA-N Ile-Asn-Phe Chemical compound CC[C@H](C)[C@H](N)C(=O)N[C@@H](CC(N)=O)C(=O)N[C@H](C(O)=O)CC1=CC=CC=C1 ZZHGKECPZXPXJF-PCBIJLKTSA-N 0.000 description 1
- HDODQNPMSHDXJT-GHCJXIJMSA-N Ile-Asn-Ser Chemical compound CC[C@H](C)[C@H](N)C(=O)N[C@@H](CC(N)=O)C(=O)N[C@@H](CO)C(O)=O HDODQNPMSHDXJT-GHCJXIJMSA-N 0.000 description 1
- QIHJTGSVGIPHIW-QSFUFRPTSA-N Ile-Asn-Val Chemical compound CC[C@H](C)[C@@H](C(=O)N[C@@H](CC(=O)N)C(=O)N[C@@H](C(C)C)C(=O)O)N QIHJTGSVGIPHIW-QSFUFRPTSA-N 0.000 description 1
- NKRJALPCDNXULF-BYULHYEWSA-N Ile-Asp-Gly Chemical compound [H]N[C@@H]([C@@H](C)CC)C(=O)N[C@@H](CC(O)=O)C(=O)NCC(O)=O NKRJALPCDNXULF-BYULHYEWSA-N 0.000 description 1
- QSPLUJGYOPZINY-ZPFDUUQYSA-N Ile-Asp-Lys Chemical compound CC[C@H](C)[C@@H](C(=O)N[C@@H](CC(=O)O)C(=O)N[C@@H](CCCCN)C(=O)O)N QSPLUJGYOPZINY-ZPFDUUQYSA-N 0.000 description 1
- DMZOUKXXHJQPTL-GRLWGSQLSA-N Ile-Gln-Ile Chemical compound CC[C@H](C)[C@@H](C(=O)N[C@@H](CCC(=O)N)C(=O)N[C@@H]([C@@H](C)CC)C(=O)O)N DMZOUKXXHJQPTL-GRLWGSQLSA-N 0.000 description 1
- BEWFWZRGBDVXRP-PEFMBERDSA-N Ile-Glu-Asn Chemical compound [H]N[C@@H]([C@@H](C)CC)C(=O)N[C@@H](CCC(O)=O)C(=O)N[C@@H](CC(N)=O)C(O)=O BEWFWZRGBDVXRP-PEFMBERDSA-N 0.000 description 1
- MTFVYKQRLXYAQN-LAEOZQHASA-N Ile-Glu-Gly Chemical compound [H]N[C@@H]([C@@H](C)CC)C(=O)N[C@@H](CCC(O)=O)C(=O)NCC(O)=O MTFVYKQRLXYAQN-LAEOZQHASA-N 0.000 description 1
- UAQSZXGJGLHMNV-XEGUGMAKSA-N Ile-Gly-Tyr Chemical compound CC[C@H](C)[C@@H](C(=O)NCC(=O)N[C@@H](CC1=CC=C(C=C1)O)C(=O)O)N UAQSZXGJGLHMNV-XEGUGMAKSA-N 0.000 description 1
- PWDSHAAAFXISLE-SXTJYALSSA-N Ile-Ile-Asp Chemical compound CC[C@H](C)[C@H](N)C(=O)N[C@@H]([C@@H](C)CC)C(=O)N[C@@H](CC(O)=O)C(O)=O PWDSHAAAFXISLE-SXTJYALSSA-N 0.000 description 1
- UWLHDGMRWXHFFY-HPCHECBXSA-N Ile-Ile-Pro Chemical compound CC[C@H](C)[C@@H](C(=O)N[C@@H]([C@@H](C)CC)C(=O)N1CCC[C@@H]1C(=O)O)N UWLHDGMRWXHFFY-HPCHECBXSA-N 0.000 description 1
- GVNNAHIRSDRIII-AJNGGQMLSA-N Ile-Lys-Lys Chemical compound CC[C@H](C)[C@@H](C(=O)N[C@@H](CCCCN)C(=O)N[C@@H](CCCCN)C(=O)O)N GVNNAHIRSDRIII-AJNGGQMLSA-N 0.000 description 1
- NLZVTPYXYXMCIP-XUXIUFHCSA-N Ile-Pro-Lys Chemical compound CC[C@H](C)[C@H](N)C(=O)N1CCC[C@H]1C(=O)N[C@@H](CCCCN)C(O)=O NLZVTPYXYXMCIP-XUXIUFHCSA-N 0.000 description 1
- ZLFNNVATRMCAKN-ZKWXMUAHSA-N Ile-Ser-Gly Chemical compound CC[C@H](C)[C@@H](C(=O)N[C@@H](CO)C(=O)NCC(=O)O)N ZLFNNVATRMCAKN-ZKWXMUAHSA-N 0.000 description 1
- YWCJXQKATPNPOE-UKJIMTQDSA-N Ile-Val-Glu Chemical compound CC[C@H](C)[C@@H](C(=O)N[C@@H](C(C)C)C(=O)N[C@@H](CCC(=O)O)C(=O)O)N YWCJXQKATPNPOE-UKJIMTQDSA-N 0.000 description 1
- 208000029462 Immunodeficiency disease Diseases 0.000 description 1
- 241001500351 Influenzavirus A Species 0.000 description 1
- 241001500350 Influenzavirus B Species 0.000 description 1
- 238000002768 Kirby-Bauer method Methods 0.000 description 1
- LZDNBBYBDGBADK-UHFFFAOYSA-N L-valyl-L-tryptophan Natural products C1=CC=C2C(CC(NC(=O)C(N)C(C)C)C(O)=O)=CNC2=C1 LZDNBBYBDGBADK-UHFFFAOYSA-N 0.000 description 1
- 102000004856 Lectins Human genes 0.000 description 1
- 108090001090 Lectins Proteins 0.000 description 1
- QPRQGENIBFLVEB-BJDJZHNGSA-N Leu-Ala-Ile Chemical compound [H]N[C@@H](CC(C)C)C(=O)N[C@@H](C)C(=O)N[C@@H]([C@@H](C)CC)C(O)=O QPRQGENIBFLVEB-BJDJZHNGSA-N 0.000 description 1
- XBBKIIGCUMBKCO-JXUBOQSCSA-N Leu-Ala-Thr Chemical compound [H]N[C@@H](CC(C)C)C(=O)N[C@@H](C)C(=O)N[C@@H]([C@@H](C)O)C(O)=O XBBKIIGCUMBKCO-JXUBOQSCSA-N 0.000 description 1
- IGUOAYLTQJLPPD-DCAQKATOSA-N Leu-Asn-Arg Chemical compound CC(C)C[C@H](N)C(=O)N[C@@H](CC(N)=O)C(=O)N[C@H](C(O)=O)CCCN=C(N)N IGUOAYLTQJLPPD-DCAQKATOSA-N 0.000 description 1
- RFUBXQQFJFGJFV-GUBZILKMSA-N Leu-Asn-Gln Chemical compound [H]N[C@@H](CC(C)C)C(=O)N[C@@H](CC(N)=O)C(=O)N[C@@H](CCC(N)=O)C(O)=O RFUBXQQFJFGJFV-GUBZILKMSA-N 0.000 description 1
- OIARJGNVARWKFP-YUMQZZPRSA-N Leu-Asn-Gly Chemical compound [H]N[C@@H](CC(C)C)C(=O)N[C@@H](CC(N)=O)C(=O)NCC(O)=O OIARJGNVARWKFP-YUMQZZPRSA-N 0.000 description 1
- POJPZSMTTMLSTG-SRVKXCTJSA-N Leu-Asn-Lys Chemical compound CC(C)C[C@@H](C(=O)N[C@@H](CC(=O)N)C(=O)N[C@@H](CCCCN)C(=O)O)N POJPZSMTTMLSTG-SRVKXCTJSA-N 0.000 description 1
- RRSLQOLASISYTB-CIUDSAMLSA-N Leu-Cys-Asp Chemical compound [H]N[C@@H](CC(C)C)C(=O)N[C@@H](CS)C(=O)N[C@@H](CC(O)=O)C(O)=O RRSLQOLASISYTB-CIUDSAMLSA-N 0.000 description 1
- PIHFVNPEAHFNLN-KKUMJFAQSA-N Leu-Cys-Tyr Chemical compound CC(C)C[C@@H](C(=O)N[C@@H](CS)C(=O)N[C@@H](CC1=CC=C(C=C1)O)C(=O)O)N PIHFVNPEAHFNLN-KKUMJFAQSA-N 0.000 description 1
- DXYBNWJZJVSZAE-GUBZILKMSA-N Leu-Gln-Cys Chemical compound CC(C)C[C@@H](C(=O)N[C@@H](CCC(=O)N)C(=O)N[C@@H](CS)C(=O)O)N DXYBNWJZJVSZAE-GUBZILKMSA-N 0.000 description 1
- HQUXQAMSWFIRET-AVGNSLFASA-N Leu-Glu-Lys Chemical compound CC(C)C[C@H](N)C(=O)N[C@@H](CCC(O)=O)C(=O)N[C@H](C(O)=O)CCCCN HQUXQAMSWFIRET-AVGNSLFASA-N 0.000 description 1
- WQWSMEOYXJTFRU-GUBZILKMSA-N Leu-Glu-Ser Chemical compound [H]N[C@@H](CC(C)C)C(=O)N[C@@H](CCC(O)=O)C(=O)N[C@@H](CO)C(O)=O WQWSMEOYXJTFRU-GUBZILKMSA-N 0.000 description 1
- LLBQJYDYOLIQAI-JYJNAYRXSA-N Leu-Glu-Tyr Chemical compound [H]N[C@@H](CC(C)C)C(=O)N[C@@H](CCC(O)=O)C(=O)N[C@@H](CC1=CC=C(O)C=C1)C(O)=O LLBQJYDYOLIQAI-JYJNAYRXSA-N 0.000 description 1
- KUIDCYNIEJBZBU-AJNGGQMLSA-N Leu-Ile-Leu Chemical compound [H]N[C@@H](CC(C)C)C(=O)N[C@@H]([C@@H](C)CC)C(=O)N[C@@H](CC(C)C)C(O)=O KUIDCYNIEJBZBU-AJNGGQMLSA-N 0.000 description 1
- REPBGZHJKYWFMJ-KKUMJFAQSA-N Leu-Lys-His Chemical compound CC(C)C[C@@H](C(=O)N[C@@H](CCCCN)C(=O)N[C@@H](CC1=CN=CN1)C(=O)O)N REPBGZHJKYWFMJ-KKUMJFAQSA-N 0.000 description 1
- RTIRBWJPYJYTLO-MELADBBJSA-N Leu-Lys-Pro Chemical compound CC(C)C[C@@H](C(=O)N[C@@H](CCCCN)C(=O)N1CCC[C@@H]1C(=O)O)N RTIRBWJPYJYTLO-MELADBBJSA-N 0.000 description 1
- BJWKOATWNQJPSK-SRVKXCTJSA-N Leu-Met-Glu Chemical compound CC(C)C[C@@H](C(=O)N[C@@H](CCSC)C(=O)N[C@@H](CCC(=O)O)C(=O)O)N BJWKOATWNQJPSK-SRVKXCTJSA-N 0.000 description 1
- INCJJHQRZGQLFC-KBPBESRZSA-N Leu-Phe-Gly Chemical compound [H]N[C@@H](CC(C)C)C(=O)N[C@@H](CC1=CC=CC=C1)C(=O)NCC(O)=O INCJJHQRZGQLFC-KBPBESRZSA-N 0.000 description 1
- IDGZVZJLYFTXSL-DCAQKATOSA-N Leu-Ser-Arg Chemical compound CC(C)C[C@H](N)C(=O)N[C@@H](CO)C(=O)N[C@H](C(O)=O)CCCN=C(N)N IDGZVZJLYFTXSL-DCAQKATOSA-N 0.000 description 1
- MVHXGBZUJLWZOH-BJDJZHNGSA-N Leu-Ser-Ile Chemical compound [H]N[C@@H](CC(C)C)C(=O)N[C@@H](CO)C(=O)N[C@@H]([C@@H](C)CC)C(O)=O MVHXGBZUJLWZOH-BJDJZHNGSA-N 0.000 description 1
- LJBVRCDPWOJOEK-PPCPHDFISA-N Leu-Thr-Ile Chemical compound [H]N[C@@H](CC(C)C)C(=O)N[C@@H]([C@@H](C)O)C(=O)N[C@@H]([C@@H](C)CC)C(O)=O LJBVRCDPWOJOEK-PPCPHDFISA-N 0.000 description 1
- UIIMIKFNIYPDJF-WDSOQIARSA-N Leu-Trp-Met Chemical compound C1=CC=C2C(C[C@@H](C(=O)N[C@@H](CCSC)C(O)=O)NC(=O)[C@@H](N)CC(C)C)=CNC2=C1 UIIMIKFNIYPDJF-WDSOQIARSA-N 0.000 description 1
- FDBTVENULFNTAL-XQQFMLRXSA-N Leu-Val-Pro Chemical compound CC(C)C[C@@H](C(=O)N[C@@H](C(C)C)C(=O)N1CCC[C@@H]1C(=O)O)N FDBTVENULFNTAL-XQQFMLRXSA-N 0.000 description 1
- ROHFNLRQFUQHCH-UHFFFAOYSA-N Leucine Natural products CC(C)CC(N)C(O)=O ROHFNLRQFUQHCH-UHFFFAOYSA-N 0.000 description 1
- NFLFJGGKOHYZJF-BJDJZHNGSA-N Lys-Ala-Ile Chemical compound CC[C@H](C)[C@@H](C(O)=O)NC(=O)[C@H](C)NC(=O)[C@@H](N)CCCCN NFLFJGGKOHYZJF-BJDJZHNGSA-N 0.000 description 1
- UWKNTTJNVSYXPC-CIUDSAMLSA-N Lys-Ala-Ser Chemical compound OC[C@@H](C(O)=O)NC(=O)[C@H](C)NC(=O)[C@@H](N)CCCCN UWKNTTJNVSYXPC-CIUDSAMLSA-N 0.000 description 1
- GAOJCVKPIGHTGO-UWVGGRQHSA-N Lys-Arg-Gly Chemical compound [H]N[C@@H](CCCCN)C(=O)N[C@@H](CCCNC(N)=N)C(=O)NCC(O)=O GAOJCVKPIGHTGO-UWVGGRQHSA-N 0.000 description 1
- NQCJGQHHYZNUDK-DCAQKATOSA-N Lys-Arg-Ser Chemical compound NCCCC[C@H](N)C(=O)N[C@H](C(=O)N[C@@H](CO)C(O)=O)CCCN=C(N)N NQCJGQHHYZNUDK-DCAQKATOSA-N 0.000 description 1
- DGWXCIORNLWGGG-CIUDSAMLSA-N Lys-Asn-Ser Chemical compound [H]N[C@@H](CCCCN)C(=O)N[C@@H](CC(N)=O)C(=O)N[C@@H](CO)C(O)=O DGWXCIORNLWGGG-CIUDSAMLSA-N 0.000 description 1
- RDIILCRAWOSDOQ-CIUDSAMLSA-N Lys-Cys-Asp Chemical compound C(CCN)C[C@@H](C(=O)N[C@@H](CS)C(=O)N[C@@H](CC(=O)O)C(=O)O)N RDIILCRAWOSDOQ-CIUDSAMLSA-N 0.000 description 1
- MLLKLNYPZRDIQG-GUBZILKMSA-N Lys-Cys-Gln Chemical compound C(CCN)C[C@@H](C(=O)N[C@@H](CS)C(=O)N[C@@H](CCC(=O)N)C(=O)O)N MLLKLNYPZRDIQG-GUBZILKMSA-N 0.000 description 1
- CKSBRMUOQDNPKZ-SRVKXCTJSA-N Lys-Gln-Met Chemical compound [H]N[C@@H](CCCCN)C(=O)N[C@@H](CCC(N)=O)C(=O)N[C@@H](CCSC)C(O)=O CKSBRMUOQDNPKZ-SRVKXCTJSA-N 0.000 description 1
- OJDFAABAHBPVTH-MNXVOIDGSA-N Lys-Ile-Gln Chemical compound [H]N[C@@H](CCCCN)C(=O)N[C@@H]([C@@H](C)CC)C(=O)N[C@@H](CCC(N)=O)C(O)=O OJDFAABAHBPVTH-MNXVOIDGSA-N 0.000 description 1
- WAIHHELKYSFIQN-XUXIUFHCSA-N Lys-Ile-Val Chemical compound [H]N[C@@H](CCCCN)C(=O)N[C@@H]([C@@H](C)CC)C(=O)N[C@@H](C(C)C)C(O)=O WAIHHELKYSFIQN-XUXIUFHCSA-N 0.000 description 1
- VUTWYNQUSJWBHO-BZSNNMDCSA-N Lys-Leu-Tyr Chemical compound [H]N[C@@H](CCCCN)C(=O)N[C@@H](CC(C)C)C(=O)N[C@@H](CC1=CC=C(O)C=C1)C(O)=O VUTWYNQUSJWBHO-BZSNNMDCSA-N 0.000 description 1
- YUAXTFMFMOIMAM-QWRGUYRKSA-N Lys-Lys-Gly Chemical compound [H]N[C@@H](CCCCN)C(=O)N[C@@H](CCCCN)C(=O)NCC(O)=O YUAXTFMFMOIMAM-QWRGUYRKSA-N 0.000 description 1
- GOVDTWNJCBRRBJ-DCAQKATOSA-N Lys-Met-Asn Chemical compound CSCC[C@@H](C(=O)N[C@@H](CC(=O)N)C(=O)O)NC(=O)[C@H](CCCCN)N GOVDTWNJCBRRBJ-DCAQKATOSA-N 0.000 description 1
- OBZHNHBAAVEWKI-DCAQKATOSA-N Lys-Pro-Asn Chemical compound NCCCC[C@H](N)C(=O)N1CCC[C@H]1C(=O)N[C@@H](CC(N)=O)C(O)=O OBZHNHBAAVEWKI-DCAQKATOSA-N 0.000 description 1
- MGKFCQFVPKOWOL-CIUDSAMLSA-N Lys-Ser-Asp Chemical compound C(CCN)C[C@@H](C(=O)N[C@@H](CO)C(=O)N[C@@H](CC(=O)O)C(=O)O)N MGKFCQFVPKOWOL-CIUDSAMLSA-N 0.000 description 1
- JMNRXRPBHFGXQX-GUBZILKMSA-N Lys-Ser-Glu Chemical compound NCCCC[C@H](N)C(=O)N[C@@H](CO)C(=O)N[C@H](C(O)=O)CCC(O)=O JMNRXRPBHFGXQX-GUBZILKMSA-N 0.000 description 1
- DIBZLYZXTSVGLN-CIUDSAMLSA-N Lys-Ser-Ser Chemical compound [H]N[C@@H](CCCCN)C(=O)N[C@@H](CO)C(=O)N[C@@H](CO)C(O)=O DIBZLYZXTSVGLN-CIUDSAMLSA-N 0.000 description 1
- HDNOQCZWJGGHSS-VEVYYDQMSA-N Met-Asn-Thr Chemical compound CSCC[C@H](N)C(=O)N[C@@H](CC(N)=O)C(=O)N[C@@H]([C@@H](C)O)C(O)=O HDNOQCZWJGGHSS-VEVYYDQMSA-N 0.000 description 1
- HGKJFNCLOHKEHS-FXQIFTODSA-N Met-Cys-Asp Chemical compound CSCC[C@H](N)C(=O)N[C@@H](CS)C(=O)N[C@H](C(O)=O)CC(O)=O HGKJFNCLOHKEHS-FXQIFTODSA-N 0.000 description 1
- RMHHNLKYPOOKQN-FXQIFTODSA-N Met-Cys-Ser Chemical compound [H]N[C@@H](CCSC)C(=O)N[C@@H](CS)C(=O)N[C@@H](CO)C(O)=O RMHHNLKYPOOKQN-FXQIFTODSA-N 0.000 description 1
- VZBXCMCHIHEPBL-SRVKXCTJSA-N Met-Glu-Lys Chemical compound CSCC[C@H](N)C(=O)N[C@@H](CCC(O)=O)C(=O)N[C@H](C(O)=O)CCCCN VZBXCMCHIHEPBL-SRVKXCTJSA-N 0.000 description 1
- RNAGAJXCSPDPRK-KKUMJFAQSA-N Met-Glu-Phe Chemical compound CSCC[C@H](N)C(=O)N[C@@H](CCC(O)=O)C(=O)N[C@H](C(O)=O)CC1=CC=CC=C1 RNAGAJXCSPDPRK-KKUMJFAQSA-N 0.000 description 1
- HLQWFLJOJRFXHO-CIUDSAMLSA-N Met-Glu-Ser Chemical compound [H]N[C@@H](CCSC)C(=O)N[C@@H](CCC(O)=O)C(=O)N[C@@H](CO)C(O)=O HLQWFLJOJRFXHO-CIUDSAMLSA-N 0.000 description 1
- IUYCGMNKIZDRQI-BQBZGAKWSA-N Met-Gly-Ala Chemical compound CSCC[C@H](N)C(=O)NCC(=O)N[C@@H](C)C(O)=O IUYCGMNKIZDRQI-BQBZGAKWSA-N 0.000 description 1
- SXWQMBGNFXAGAT-FJXKBIBVSA-N Met-Gly-Thr Chemical compound CSCC[C@H](N)C(=O)NCC(=O)N[C@@H]([C@@H](C)O)C(O)=O SXWQMBGNFXAGAT-FJXKBIBVSA-N 0.000 description 1
- CGUYGMFQZCYJSG-DCAQKATOSA-N Met-Lys-Ser Chemical compound [H]N[C@@H](CCSC)C(=O)N[C@@H](CCCCN)C(=O)N[C@@H](CO)C(O)=O CGUYGMFQZCYJSG-DCAQKATOSA-N 0.000 description 1
- MQASRXPTQJJNFM-JYJNAYRXSA-N Met-Pro-Phe Chemical compound CSCC[C@H](N)C(=O)N1CCC[C@H]1C(=O)N[C@H](C(O)=O)CC1=CC=CC=C1 MQASRXPTQJJNFM-JYJNAYRXSA-N 0.000 description 1
- YGNUDKAPJARTEM-GUBZILKMSA-N Met-Val-Ala Chemical compound CSCC[C@H](N)C(=O)N[C@@H](C(C)C)C(=O)N[C@@H](C)C(O)=O YGNUDKAPJARTEM-GUBZILKMSA-N 0.000 description 1
- XZFYRXDAULDNFX-UHFFFAOYSA-N N-L-cysteinyl-L-phenylalanine Natural products SCC(N)C(=O)NC(C(O)=O)CC1=CC=CC=C1 XZFYRXDAULDNFX-UHFFFAOYSA-N 0.000 description 1
- 108010002311 N-glycylglutamic acid Proteins 0.000 description 1
- 108010066427 N-valyltryptophan Proteins 0.000 description 1
- 108700001237 Nucleic Acid-Based Vaccines Proteins 0.000 description 1
- 241000712464 Orthomyxoviridae Species 0.000 description 1
- 238000012408 PCR amplification Methods 0.000 description 1
- 241001631646 Papillomaviridae Species 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
- 241000286209 Phasianidae Species 0.000 description 1
- HCTXJGRYAACKOB-SRVKXCTJSA-N Phe-Asn-Asp Chemical compound C1=CC=C(C=C1)C[C@@H](C(=O)N[C@@H](CC(=O)N)C(=O)N[C@@H](CC(=O)O)C(=O)O)N HCTXJGRYAACKOB-SRVKXCTJSA-N 0.000 description 1
- MGBRZXXGQBAULP-DRZSPHRISA-N Phe-Glu-Ala Chemical compound OC(=O)[C@H](C)NC(=O)[C@H](CCC(O)=O)NC(=O)[C@@H](N)CC1=CC=CC=C1 MGBRZXXGQBAULP-DRZSPHRISA-N 0.000 description 1
- PSKRILMFHNIUAO-JYJNAYRXSA-N Phe-Glu-Lys Chemical compound C1=CC=C(C=C1)C[C@@H](C(=O)N[C@@H](CCC(=O)O)C(=O)N[C@@H](CCCCN)C(=O)O)N PSKRILMFHNIUAO-JYJNAYRXSA-N 0.000 description 1
- OYQBFWWQSVIHBN-FHWLQOOXSA-N Phe-Glu-Phe Chemical compound [H]N[C@@H](CC1=CC=CC=C1)C(=O)N[C@@H](CCC(O)=O)C(=O)N[C@@H](CC1=CC=CC=C1)C(O)=O OYQBFWWQSVIHBN-FHWLQOOXSA-N 0.000 description 1
- JWQWPTLEOFNCGX-AVGNSLFASA-N Phe-Glu-Ser Chemical compound OC[C@@H](C(O)=O)NC(=O)[C@H](CCC(O)=O)NC(=O)[C@@H](N)CC1=CC=CC=C1 JWQWPTLEOFNCGX-AVGNSLFASA-N 0.000 description 1
- JJHVFCUWLSKADD-ONGXEEELSA-N Phe-Gly-Ala Chemical compound [H]N[C@@H](CC1=CC=CC=C1)C(=O)NCC(=O)N[C@@H](C)C(O)=O JJHVFCUWLSKADD-ONGXEEELSA-N 0.000 description 1
- VZFPYFRVHMSSNA-JURCDPSOSA-N Phe-Ile-Ala Chemical compound OC(=O)[C@H](C)NC(=O)[C@H]([C@@H](C)CC)NC(=O)[C@@H](N)CC1=CC=CC=C1 VZFPYFRVHMSSNA-JURCDPSOSA-N 0.000 description 1
- YKUGPVXSDOOANW-KKUMJFAQSA-N Phe-Leu-Asp Chemical compound [H]N[C@@H](CC1=CC=CC=C1)C(=O)N[C@@H](CC(C)C)C(=O)N[C@@H](CC(O)=O)C(O)=O YKUGPVXSDOOANW-KKUMJFAQSA-N 0.000 description 1
- OXKJSGGTHFMGDT-UFYCRDLUSA-N Phe-Phe-Arg Chemical compound C([C@H](N)C(=O)N[C@@H](CC=1C=CC=CC=1)C(=O)N[C@@H](CCCNC(N)=N)C(O)=O)C1=CC=CC=C1 OXKJSGGTHFMGDT-UFYCRDLUSA-N 0.000 description 1
- YMTMNYNEZDAGMW-RNXOBYDBSA-N Phe-Phe-Trp Chemical compound C1=CC=C(C=C1)C[C@@H](C(=O)N[C@@H](CC2=CC=CC=C2)C(=O)N[C@@H](CC3=CNC4=CC=CC=C43)C(=O)O)N YMTMNYNEZDAGMW-RNXOBYDBSA-N 0.000 description 1
- LKRUQZQZMXMKEQ-SFJXLCSZSA-N Phe-Trp-Thr Chemical compound [H]N[C@@H](CC1=CC=CC=C1)C(=O)N[C@@H](CC1=CNC2=C1C=CC=C2)C(=O)N[C@@H]([C@@H](C)O)C(O)=O LKRUQZQZMXMKEQ-SFJXLCSZSA-N 0.000 description 1
- 235000014676 Phragmites communis Nutrition 0.000 description 1
- DBALDZKOTNSBFM-FXQIFTODSA-N Pro-Ala-Asn Chemical compound [H]N1CCC[C@H]1C(=O)N[C@@H](C)C(=O)N[C@@H](CC(N)=O)C(O)=O DBALDZKOTNSBFM-FXQIFTODSA-N 0.000 description 1
- LSIWVWRUTKPXDS-DCAQKATOSA-N Pro-Gln-Arg Chemical compound [H]N1CCC[C@H]1C(=O)N[C@@H](CCC(N)=O)C(=O)N[C@@H](CCCNC(N)=N)C(O)=O LSIWVWRUTKPXDS-DCAQKATOSA-N 0.000 description 1
- KTFZQPLSPLWLKN-KKUMJFAQSA-N Pro-Gln-Tyr Chemical compound [H]N1CCC[C@H]1C(=O)N[C@@H](CCC(N)=O)C(=O)N[C@@H](CC1=CC=C(O)C=C1)C(O)=O KTFZQPLSPLWLKN-KKUMJFAQSA-N 0.000 description 1
- ZTVCLZLGHZXLOT-ULQDDVLXSA-N Pro-Glu-Trp Chemical compound C1C[C@H](NC1)C(=O)N[C@@H](CCC(=O)O)C(=O)N[C@@H](CC2=CNC3=CC=CC=C32)C(=O)O ZTVCLZLGHZXLOT-ULQDDVLXSA-N 0.000 description 1
- QGOZJLYCGRYYRW-KKUMJFAQSA-N Pro-Glu-Tyr Chemical compound [H]N1CCC[C@H]1C(=O)N[C@@H](CCC(O)=O)C(=O)N[C@@H](CC1=CC=C(O)C=C1)C(O)=O QGOZJLYCGRYYRW-KKUMJFAQSA-N 0.000 description 1
- ULIWFCCJIOEHMU-BQBZGAKWSA-N Pro-Gly-Asp Chemical compound OC(=O)C[C@@H](C(O)=O)NC(=O)CNC(=O)[C@@H]1CCCN1 ULIWFCCJIOEHMU-BQBZGAKWSA-N 0.000 description 1
- BRJGUPWVFXKBQI-XUXIUFHCSA-N Pro-Leu-Ile Chemical compound [H]N1CCC[C@H]1C(=O)N[C@@H](CC(C)C)C(=O)N[C@@H]([C@@H](C)CC)C(O)=O BRJGUPWVFXKBQI-XUXIUFHCSA-N 0.000 description 1
- WCNVGGZRTNHOOS-ULQDDVLXSA-N Pro-Lys-Tyr Chemical compound C1C[C@H](NC1)C(=O)N[C@@H](CCCCN)C(=O)N[C@@H](CC2=CC=C(C=C2)O)C(=O)O WCNVGGZRTNHOOS-ULQDDVLXSA-N 0.000 description 1
- WOIFYRZPIORBRY-AVGNSLFASA-N Pro-Lys-Val Chemical compound [H]N1CCC[C@H]1C(=O)N[C@@H](CCCCN)C(=O)N[C@@H](C(C)C)C(O)=O WOIFYRZPIORBRY-AVGNSLFASA-N 0.000 description 1
- FRVUYKWGPCQRBL-GUBZILKMSA-N Pro-Met-Cys Chemical compound CSCC[C@@H](C(=O)N[C@@H](CS)C(=O)O)NC(=O)[C@@H]1CCCN1 FRVUYKWGPCQRBL-GUBZILKMSA-N 0.000 description 1
- WIPAMEKBSHNFQE-IUCAKERBSA-N Pro-Met-Gly Chemical compound CSCC[C@@H](C(=O)NCC(=O)O)NC(=O)[C@@H]1CCCN1 WIPAMEKBSHNFQE-IUCAKERBSA-N 0.000 description 1
- GNADVDLLGVSXLS-ULQDDVLXSA-N Pro-Phe-His Chemical compound [H]N1CCC[C@H]1C(=O)N[C@@H](CC1=CC=CC=C1)C(=O)N[C@@H](CC1=CNC=N1)C(O)=O GNADVDLLGVSXLS-ULQDDVLXSA-N 0.000 description 1
- GZNYIXWOIUFLGO-ZJDVBMNYSA-N Pro-Thr-Thr Chemical compound [H]N1CCC[C@H]1C(=O)N[C@@H]([C@@H](C)O)C(=O)N[C@@H]([C@@H](C)O)C(O)=O GZNYIXWOIUFLGO-ZJDVBMNYSA-N 0.000 description 1
- ONIBWKKTOPOVIA-UHFFFAOYSA-N Proline Natural products OC(=O)C1CCCN1 ONIBWKKTOPOVIA-UHFFFAOYSA-N 0.000 description 1
- 102100035123 Retrotransposon-like protein 1 Human genes 0.000 description 1
- 241000315672 SARS coronavirus Species 0.000 description 1
- 241000242680 Schistosoma mansoni Species 0.000 description 1
- 238000012300 Sequence Analysis Methods 0.000 description 1
- BKOKTRCZXRIQPX-ZLUOBGJFSA-N Ser-Ala-Cys Chemical compound C[C@@H](C(=O)N[C@@H](CS)C(=O)O)NC(=O)[C@H](CO)N BKOKTRCZXRIQPX-ZLUOBGJFSA-N 0.000 description 1
- OYEDZGNMSBZCIM-XGEHTFHBSA-N Ser-Arg-Thr Chemical compound [H]N[C@@H](CO)C(=O)N[C@@H](CCCNC(N)=N)C(=O)N[C@@H]([C@@H](C)O)C(O)=O OYEDZGNMSBZCIM-XGEHTFHBSA-N 0.000 description 1
- ZXLUWXWISXIFIX-ACZMJKKPSA-N Ser-Asn-Glu Chemical compound [H]N[C@@H](CO)C(=O)N[C@@H](CC(N)=O)C(=O)N[C@@H](CCC(O)=O)C(O)=O ZXLUWXWISXIFIX-ACZMJKKPSA-N 0.000 description 1
- UOLGINIHBRIECN-FXQIFTODSA-N Ser-Glu-Glu Chemical compound [H]N[C@@H](CO)C(=O)N[C@@H](CCC(O)=O)C(=O)N[C@@H](CCC(O)=O)C(O)=O UOLGINIHBRIECN-FXQIFTODSA-N 0.000 description 1
- AEGUWTFAQQWVLC-BQBZGAKWSA-N Ser-Gly-Arg Chemical compound [H]N[C@@H](CO)C(=O)NCC(=O)N[C@@H](CCCNC(N)=N)C(O)=O AEGUWTFAQQWVLC-BQBZGAKWSA-N 0.000 description 1
- OQPNSDWGAMFJNU-QWRGUYRKSA-N Ser-Gly-Tyr Chemical compound OC[C@H](N)C(=O)NCC(=O)N[C@H](C(O)=O)CC1=CC=C(O)C=C1 OQPNSDWGAMFJNU-QWRGUYRKSA-N 0.000 description 1
- XXXAXOWMBOKTRN-XPUUQOCRSA-N Ser-Gly-Val Chemical compound [H]N[C@@H](CO)C(=O)NCC(=O)N[C@@H](C(C)C)C(O)=O XXXAXOWMBOKTRN-XPUUQOCRSA-N 0.000 description 1
- MQQBBLVOUUJKLH-HJPIBITLSA-N Ser-Ile-Tyr Chemical compound [H]N[C@@H](CO)C(=O)N[C@@H]([C@@H](C)CC)C(=O)N[C@@H](CC1=CC=C(O)C=C1)C(O)=O MQQBBLVOUUJKLH-HJPIBITLSA-N 0.000 description 1
- GJFYFGOEWLDQGW-GUBZILKMSA-N Ser-Leu-Gln Chemical compound CC(C)C[C@@H](C(=O)N[C@@H](CCC(=O)N)C(=O)O)NC(=O)[C@H](CO)N GJFYFGOEWLDQGW-GUBZILKMSA-N 0.000 description 1
- GVIGVIOEYBOTCB-XIRDDKMYSA-N Ser-Leu-Trp Chemical compound C1=CC=C2C(C[C@H](NC(=O)[C@@H](NC(=O)[C@@H](N)CO)CC(C)C)C(O)=O)=CNC2=C1 GVIGVIOEYBOTCB-XIRDDKMYSA-N 0.000 description 1
- IXZHZUGGKLRHJD-DCAQKATOSA-N Ser-Leu-Val Chemical compound [H]N[C@@H](CO)C(=O)N[C@@H](CC(C)C)C(=O)N[C@@H](C(C)C)C(O)=O IXZHZUGGKLRHJD-DCAQKATOSA-N 0.000 description 1
- ADJDNJCSPNFFPI-FXQIFTODSA-N Ser-Pro-Ala Chemical compound OC(=O)[C@H](C)NC(=O)[C@@H]1CCCN1C(=O)[C@@H](N)CO ADJDNJCSPNFFPI-FXQIFTODSA-N 0.000 description 1
- HHJFMHQYEAAOBM-ZLUOBGJFSA-N Ser-Ser-Ala Chemical compound [H]N[C@@H](CO)C(=O)N[C@@H](CO)C(=O)N[C@@H](C)C(O)=O HHJFMHQYEAAOBM-ZLUOBGJFSA-N 0.000 description 1
- AABIBDJHSKIMJK-FXQIFTODSA-N Ser-Ser-Met Chemical compound [H]N[C@@H](CO)C(=O)N[C@@H](CO)C(=O)N[C@@H](CCSC)C(O)=O AABIBDJHSKIMJK-FXQIFTODSA-N 0.000 description 1
- ILZAUMFXKSIUEF-SRVKXCTJSA-N Ser-Ser-Phe Chemical compound OC[C@H](N)C(=O)N[C@@H](CO)C(=O)N[C@H](C(O)=O)CC1=CC=CC=C1 ILZAUMFXKSIUEF-SRVKXCTJSA-N 0.000 description 1
- SZRNDHWMVSFPSP-XKBZYTNZSA-N Ser-Thr-Gln Chemical compound C[C@H]([C@@H](C(=O)N[C@@H](CCC(=O)N)C(=O)O)NC(=O)[C@H](CO)N)O SZRNDHWMVSFPSP-XKBZYTNZSA-N 0.000 description 1
- KKKVOZNCLALMPV-XKBZYTNZSA-N Ser-Thr-Glu Chemical compound [H]N[C@@H](CO)C(=O)N[C@@H]([C@@H](C)O)C(=O)N[C@@H](CCC(O)=O)C(O)=O KKKVOZNCLALMPV-XKBZYTNZSA-N 0.000 description 1
- BDMWLJLPPUCLNV-XGEHTFHBSA-N Ser-Thr-Val Chemical compound [H]N[C@@H](CO)C(=O)N[C@@H]([C@@H](C)O)C(=O)N[C@@H](C(C)C)C(O)=O BDMWLJLPPUCLNV-XGEHTFHBSA-N 0.000 description 1
- ZWSZBWAFDZRBNM-UBHSHLNASA-N Ser-Trp-Ser Chemical compound [H]N[C@@H](CO)C(=O)N[C@@H](CC1=CNC2=C1C=CC=C2)C(=O)N[C@@H](CO)C(O)=O ZWSZBWAFDZRBNM-UBHSHLNASA-N 0.000 description 1
- UBTNVMGPMYDYIU-HJPIBITLSA-N Ser-Tyr-Ile Chemical compound [H]N[C@@H](CO)C(=O)N[C@@H](CC1=CC=C(O)C=C1)C(=O)N[C@@H]([C@@H](C)CC)C(O)=O UBTNVMGPMYDYIU-HJPIBITLSA-N 0.000 description 1
- PMTWIUBUQRGCSB-FXQIFTODSA-N Ser-Val-Ala Chemical compound [H]N[C@@H](CO)C(=O)N[C@@H](C(C)C)C(=O)N[C@@H](C)C(O)=O PMTWIUBUQRGCSB-FXQIFTODSA-N 0.000 description 1
- JZRYFUGREMECBH-XPUUQOCRSA-N Ser-Val-Gly Chemical compound [H]N[C@@H](CO)C(=O)N[C@@H](C(C)C)C(=O)NCC(O)=O JZRYFUGREMECBH-XPUUQOCRSA-N 0.000 description 1
- LGIMRDKGABDMBN-DCAQKATOSA-N Ser-Val-Lys Chemical compound CC(C)[C@@H](C(=O)N[C@@H](CCCCN)C(=O)O)NC(=O)[C@H](CO)N LGIMRDKGABDMBN-DCAQKATOSA-N 0.000 description 1
- MTCFGRXMJLQNBG-UHFFFAOYSA-N Serine Natural products OCC(N)C(O)=O MTCFGRXMJLQNBG-UHFFFAOYSA-N 0.000 description 1
- 108091081024 Start codon Proteins 0.000 description 1
- 101710172711 Structural protein Proteins 0.000 description 1
- GZYNMZQXFRWDFH-YTWAJWBKSA-N Thr-Arg-Pro Chemical compound C[C@H]([C@@H](C(=O)N[C@@H](CCCN=C(N)N)C(=O)N1CCC[C@@H]1C(=O)O)N)O GZYNMZQXFRWDFH-YTWAJWBKSA-N 0.000 description 1
- JHBHMCMKSPXRHV-NUMRIWBASA-N Thr-Asn-Gln Chemical compound C[C@H]([C@@H](C(=O)N[C@@H](CC(=O)N)C(=O)N[C@@H](CCC(=O)N)C(=O)O)N)O JHBHMCMKSPXRHV-NUMRIWBASA-N 0.000 description 1
- PAOYNIKMYOGBMR-PBCZWWQYSA-N Thr-Asn-His Chemical compound C[C@H]([C@@H](C(=O)N[C@@H](CC(=O)N)C(=O)N[C@@H](CC1=CN=CN1)C(=O)O)N)O PAOYNIKMYOGBMR-PBCZWWQYSA-N 0.000 description 1
- XXNLGZRRSKPSGF-HTUGSXCWSA-N Thr-Gln-Phe Chemical compound C[C@H]([C@@H](C(=O)N[C@@H](CCC(=O)N)C(=O)N[C@@H](CC1=CC=CC=C1)C(=O)O)N)O XXNLGZRRSKPSGF-HTUGSXCWSA-N 0.000 description 1
- GKWNLDNXMMLRMC-GLLZPBPUSA-N Thr-Glu-Gln Chemical compound C[C@H]([C@@H](C(=O)N[C@@H](CCC(=O)O)C(=O)N[C@@H](CCC(=O)N)C(=O)O)N)O GKWNLDNXMMLRMC-GLLZPBPUSA-N 0.000 description 1
- VUSAEKOXGNEYNE-PBCZWWQYSA-N Thr-His-Asn Chemical compound C[C@H]([C@@H](C(=O)N[C@@H](CC1=CN=CN1)C(=O)N[C@@H](CC(=O)N)C(=O)O)N)O VUSAEKOXGNEYNE-PBCZWWQYSA-N 0.000 description 1
- URPSJRMWHQTARR-MBLNEYKQSA-N Thr-Ile-Gly Chemical compound [H]N[C@@H]([C@@H](C)O)C(=O)N[C@@H]([C@@H](C)CC)C(=O)NCC(O)=O URPSJRMWHQTARR-MBLNEYKQSA-N 0.000 description 1
- ADPHPKGWVDHWML-PPCPHDFISA-N Thr-Ile-Leu Chemical compound CC[C@H](C)[C@@H](C(=O)N[C@@H](CC(C)C)C(=O)O)NC(=O)[C@H]([C@@H](C)O)N ADPHPKGWVDHWML-PPCPHDFISA-N 0.000 description 1
- AHOLTQCAVBSUDP-PPCPHDFISA-N Thr-Ile-Lys Chemical compound CC[C@H](C)[C@H](NC(=O)[C@@H](N)[C@@H](C)O)C(=O)N[C@@H](CCCCN)C(O)=O AHOLTQCAVBSUDP-PPCPHDFISA-N 0.000 description 1
- LCCSEJSPBWKBNT-OSUNSFLBSA-N Thr-Ile-Met Chemical compound CC[C@H](C)[C@@H](C(=O)N[C@@H](CCSC)C(=O)O)NC(=O)[C@H]([C@@H](C)O)N LCCSEJSPBWKBNT-OSUNSFLBSA-N 0.000 description 1
- IMDMLDSVUSMAEJ-HJGDQZAQSA-N Thr-Leu-Asn Chemical compound [H]N[C@@H]([C@@H](C)O)C(=O)N[C@@H](CC(C)C)C(=O)N[C@@H](CC(N)=O)C(O)=O IMDMLDSVUSMAEJ-HJGDQZAQSA-N 0.000 description 1
- RRRRCRYTLZVCEN-HJGDQZAQSA-N Thr-Leu-Asp Chemical compound [H]N[C@@H]([C@@H](C)O)C(=O)N[C@@H](CC(C)C)C(=O)N[C@@H](CC(O)=O)C(O)=O RRRRCRYTLZVCEN-HJGDQZAQSA-N 0.000 description 1
- HPQHHRLWSAMMKG-KATARQTJSA-N Thr-Lys-Cys Chemical compound C[C@H]([C@@H](C(=O)N[C@@H](CCCCN)C(=O)N[C@@H](CS)C(=O)O)N)O HPQHHRLWSAMMKG-KATARQTJSA-N 0.000 description 1
- SPVHQURZJCUDQC-VOAKCMCISA-N Thr-Lys-Leu Chemical compound [H]N[C@@H]([C@@H](C)O)C(=O)N[C@@H](CCCCN)C(=O)N[C@@H](CC(C)C)C(O)=O SPVHQURZJCUDQC-VOAKCMCISA-N 0.000 description 1
- RVMNUBQWPVOUKH-HEIBUPTGSA-N Thr-Ser-Thr Chemical compound [H]N[C@@H]([C@@H](C)O)C(=O)N[C@@H](CO)C(=O)N[C@@H]([C@@H](C)O)C(O)=O RVMNUBQWPVOUKH-HEIBUPTGSA-N 0.000 description 1
- RPECVQBNONKZAT-WZLNRYEVSA-N Thr-Tyr-Ile Chemical compound CC[C@H](C)[C@@H](C(=O)O)NC(=O)[C@H](CC1=CC=C(C=C1)O)NC(=O)[C@H]([C@@H](C)O)N RPECVQBNONKZAT-WZLNRYEVSA-N 0.000 description 1
- KZTLZZQTJMCGIP-ZJDVBMNYSA-N Thr-Val-Thr Chemical compound [H]N[C@@H]([C@@H](C)O)C(=O)N[C@@H](C(C)C)C(=O)N[C@@H]([C@@H](C)O)C(O)=O KZTLZZQTJMCGIP-ZJDVBMNYSA-N 0.000 description 1
- AYFVYJQAPQTCCC-UHFFFAOYSA-N Threonine Natural products CC(O)C(N)C(O)=O AYFVYJQAPQTCCC-UHFFFAOYSA-N 0.000 description 1
- 239000004473 Threonine Substances 0.000 description 1
- WKCFCVBOFKEVKY-HSCHXYMDSA-N Trp-Leu-Ile Chemical compound CC[C@H](C)[C@@H](C(=O)O)NC(=O)[C@H](CC(C)C)NC(=O)[C@H](CC1=CNC2=CC=CC=C21)N WKCFCVBOFKEVKY-HSCHXYMDSA-N 0.000 description 1
- CCZXBOFIBYQLEV-IHPCNDPISA-N Trp-Leu-Leu Chemical compound CC(C)C[C@H](NC(=O)[C@H](CC(C)C)NC(=O)[C@@H](N)Cc1c[nH]c2ccccc12)C(O)=O CCZXBOFIBYQLEV-IHPCNDPISA-N 0.000 description 1
- KBKTUNYBNJWFRL-UBHSHLNASA-N Trp-Ser-Asn Chemical compound C1=CC=C2C(C[C@H](N)C(=O)N[C@@H](CO)C(=O)N[C@@H](CC(N)=O)C(O)=O)=CNC2=C1 KBKTUNYBNJWFRL-UBHSHLNASA-N 0.000 description 1
- ABRICLFKFRFDKS-IHPCNDPISA-N Trp-Ser-Tyr Chemical compound C([C@H](NC(=O)[C@H](CO)NC(=O)[C@H](CC=1C2=CC=CC=C2NC=1)N)C(O)=O)C1=CC=C(O)C=C1 ABRICLFKFRFDKS-IHPCNDPISA-N 0.000 description 1
- ZSSKZJBTPJBKFT-WZUXKDABSA-N Trp-Tyr-Gly-Tyr-His-His Chemical compound N[C@@H](Cc1c[nH]c2ccccc12)C(=O)N[C@@H](Cc1ccc(O)cc1)C(=O)NCC(=O)N[C@@H](Cc1ccc(O)cc1)C(=O)N[C@@H](Cc1cnc[nH]1)C(=O)N[C@@H](Cc1cnc[nH]1)C(O)=O ZSSKZJBTPJBKFT-WZUXKDABSA-N 0.000 description 1
- QIVBCDIJIAJPQS-UHFFFAOYSA-N Tryptophan Natural products C1=CC=C2C(CC(N)C(O)=O)=CNC2=C1 QIVBCDIJIAJPQS-UHFFFAOYSA-N 0.000 description 1
- VCXWRWYFJLXITF-AUTRQRHGSA-N Tyr-Ala-Ala Chemical compound OC(=O)[C@H](C)NC(=O)[C@H](C)NC(=O)[C@@H](N)CC1=CC=C(O)C=C1 VCXWRWYFJLXITF-AUTRQRHGSA-N 0.000 description 1
- KSVMDJJCYKIXTK-IGNZVWTISA-N Tyr-Ala-Tyr Chemical compound C([C@H](N)C(=O)N[C@@H](C)C(=O)N[C@@H](CC=1C=CC(O)=CC=1)C(O)=O)C1=CC=C(O)C=C1 KSVMDJJCYKIXTK-IGNZVWTISA-N 0.000 description 1
- PZXUIGWOEWWFQM-SRVKXCTJSA-N Tyr-Asn-Asn Chemical compound [H]N[C@@H](CC1=CC=C(O)C=C1)C(=O)N[C@@H](CC(N)=O)C(=O)N[C@@H](CC(N)=O)C(O)=O PZXUIGWOEWWFQM-SRVKXCTJSA-N 0.000 description 1
- JFDGVHXRCKEBAU-KKUMJFAQSA-N Tyr-Asp-Lys Chemical compound C1=CC(=CC=C1C[C@@H](C(=O)N[C@@H](CC(=O)O)C(=O)N[C@@H](CCCCN)C(=O)O)N)O JFDGVHXRCKEBAU-KKUMJFAQSA-N 0.000 description 1
- TZXFLDNBYYGLKA-BZSNNMDCSA-N Tyr-Asp-Tyr Chemical compound C([C@H](N)C(=O)N[C@@H](CC(O)=O)C(=O)N[C@@H](CC=1C=CC(O)=CC=1)C(O)=O)C1=CC=C(O)C=C1 TZXFLDNBYYGLKA-BZSNNMDCSA-N 0.000 description 1
- CRHFOYCJGVJPLE-AVGNSLFASA-N Tyr-Gln-Asn Chemical compound C1=CC(=CC=C1C[C@@H](C(=O)N[C@@H](CCC(=O)N)C(=O)N[C@@H](CC(=O)N)C(=O)O)N)O CRHFOYCJGVJPLE-AVGNSLFASA-N 0.000 description 1
- UXUFNBVCPAWACG-SIUGBPQLSA-N Tyr-Gln-Ile Chemical compound CC[C@H](C)[C@@H](C(=O)O)NC(=O)[C@H](CCC(=O)N)NC(=O)[C@H](CC1=CC=C(C=C1)O)N UXUFNBVCPAWACG-SIUGBPQLSA-N 0.000 description 1
- WVRUKYLYMFGKAN-IHRRRGAJSA-N Tyr-Glu-Glu Chemical compound OC(=O)CC[C@@H](C(O)=O)NC(=O)[C@H](CCC(O)=O)NC(=O)[C@@H](N)CC1=CC=C(O)C=C1 WVRUKYLYMFGKAN-IHRRRGAJSA-N 0.000 description 1
- CDHQEOXPWBDFPL-QWRGUYRKSA-N Tyr-Gly-Asn Chemical compound NC(=O)C[C@@H](C(O)=O)NC(=O)CNC(=O)[C@@H](N)CC1=CC=C(O)C=C1 CDHQEOXPWBDFPL-QWRGUYRKSA-N 0.000 description 1
- KEANSLVUGJADPN-LKTVYLICSA-N Tyr-His-Ala Chemical compound C[C@@H](C(=O)O)NC(=O)[C@H](CC1=CN=CN1)NC(=O)[C@H](CC2=CC=C(C=C2)O)N KEANSLVUGJADPN-LKTVYLICSA-N 0.000 description 1
- MVYRJYISVJWKSX-KBPBESRZSA-N Tyr-His-Gly Chemical compound C1=CC(=CC=C1C[C@@H](C(=O)N[C@@H](CC2=CN=CN2)C(=O)NCC(=O)O)N)O MVYRJYISVJWKSX-KBPBESRZSA-N 0.000 description 1
- WVGKPKDWYQXWLU-BZSNNMDCSA-N Tyr-His-Lys Chemical compound C1=CC(=CC=C1C[C@@H](C(=O)N[C@@H](CC2=CN=CN2)C(=O)N[C@@H](CCCCN)C(=O)O)N)O WVGKPKDWYQXWLU-BZSNNMDCSA-N 0.000 description 1
- QARCDOCCDOLJSF-HJPIBITLSA-N Tyr-Ile-Ser Chemical compound CC[C@H](C)[C@@H](C(=O)N[C@@H](CO)C(=O)O)NC(=O)[C@H](CC1=CC=C(C=C1)O)N QARCDOCCDOLJSF-HJPIBITLSA-N 0.000 description 1
- ARMNWLJYHCOSHE-KKUMJFAQSA-N Tyr-Pro-Gln Chemical compound [H]N[C@@H](CC1=CC=C(O)C=C1)C(=O)N1CCC[C@H]1C(=O)N[C@@H](CCC(N)=O)C(O)=O ARMNWLJYHCOSHE-KKUMJFAQSA-N 0.000 description 1
- XGZBEGGGAUQBMB-KJEVXHAQSA-N Tyr-Pro-Thr Chemical compound C[C@H]([C@@H](C(=O)O)NC(=O)[C@@H]1CCCN1C(=O)[C@H](CC2=CC=C(C=C2)O)N)O XGZBEGGGAUQBMB-KJEVXHAQSA-N 0.000 description 1
- QFXVAFIHVWXXBJ-AVGNSLFASA-N Tyr-Ser-Glu Chemical compound [H]N[C@@H](CC1=CC=C(O)C=C1)C(=O)N[C@@H](CO)C(=O)N[C@@H](CCC(O)=O)C(O)=O QFXVAFIHVWXXBJ-AVGNSLFASA-N 0.000 description 1
- ASQFIHTXXMFENG-XPUUQOCRSA-N Val-Ala-Gly Chemical compound CC(C)[C@H](N)C(=O)N[C@@H](C)C(=O)NCC(O)=O ASQFIHTXXMFENG-XPUUQOCRSA-N 0.000 description 1
- QHDXUYOYTPWCSK-RCOVLWMOSA-N Val-Asp-Gly Chemical compound CC(C)[C@@H](C(=O)N[C@@H](CC(=O)O)C(=O)NCC(=O)O)N QHDXUYOYTPWCSK-RCOVLWMOSA-N 0.000 description 1
- JTWIMNMUYLQNPI-WPRPVWTQSA-N Val-Gly-Arg Chemical compound CC(C)[C@H](N)C(=O)NCC(=O)N[C@H](C(O)=O)CCCNC(N)=N JTWIMNMUYLQNPI-WPRPVWTQSA-N 0.000 description 1
- ZRSZTKTVPNSUNA-IHRRRGAJSA-N Val-Lys-Leu Chemical compound CC(C)C[C@H](NC(=O)[C@H](CCCCN)NC(=O)[C@@H](N)C(C)C)C(O)=O ZRSZTKTVPNSUNA-IHRRRGAJSA-N 0.000 description 1
- YMTOEGGOCHVGEH-IHRRRGAJSA-N Val-Lys-Lys Chemical compound CC(C)[C@H](N)C(=O)N[C@@H](CCCCN)C(=O)N[C@@H](CCCCN)C(O)=O YMTOEGGOCHVGEH-IHRRRGAJSA-N 0.000 description 1
- CXWJFWAZIVWBOS-XQQFMLRXSA-N Val-Lys-Pro Chemical compound CC(C)[C@@H](C(=O)N[C@@H](CCCCN)C(=O)N1CCC[C@@H]1C(=O)O)N CXWJFWAZIVWBOS-XQQFMLRXSA-N 0.000 description 1
- JAKHAONCJJZVHT-DCAQKATOSA-N Val-Lys-Ser Chemical compound CC(C)[C@@H](C(=O)N[C@@H](CCCCN)C(=O)N[C@@H](CO)C(=O)O)N JAKHAONCJJZVHT-DCAQKATOSA-N 0.000 description 1
- PQSNETRGCRUOGP-KKHAAJSZSA-N Val-Thr-Asn Chemical compound CC(C)[C@H](N)C(=O)N[C@@H]([C@@H](C)O)C(=O)N[C@H](C(O)=O)CC(N)=O PQSNETRGCRUOGP-KKHAAJSZSA-N 0.000 description 1
- TVGWMCTYUFBXAP-QTKMDUPCSA-N Val-Thr-His Chemical compound C[C@H]([C@@H](C(=O)N[C@@H](CC1=CN=CN1)C(=O)O)NC(=O)[C@H](C(C)C)N)O TVGWMCTYUFBXAP-QTKMDUPCSA-N 0.000 description 1
- ZLMFVXMJFIWIRE-FHWLQOOXSA-N Val-Trp-Leu Chemical compound CC(C)C[C@@H](C(=O)O)NC(=O)[C@H](CC1=CNC2=CC=CC=C21)NC(=O)[C@H](C(C)C)N ZLMFVXMJFIWIRE-FHWLQOOXSA-N 0.000 description 1
- OEVFFOBAXHBXKM-HSHDSVGOSA-N Val-Trp-Thr Chemical compound C[C@H]([C@@H](C(=O)O)NC(=O)[C@H](CC1=CNC2=CC=CC=C21)NC(=O)[C@H](C(C)C)N)O OEVFFOBAXHBXKM-HSHDSVGOSA-N 0.000 description 1
- 208000036142 Viral infection Diseases 0.000 description 1
- JLCPHMBAVCMARE-UHFFFAOYSA-N [3-[[3-[[3-[[3-[[3-[[3-[[3-[[3-[[3-[[3-[[3-[[5-(2-amino-6-oxo-1H-purin-9-yl)-3-[[3-[[3-[[3-[[3-[[3-[[5-(2-amino-6-oxo-1H-purin-9-yl)-3-[[5-(2-amino-6-oxo-1H-purin-9-yl)-3-hydroxyoxolan-2-yl]methoxy-hydroxyphosphoryl]oxyoxolan-2-yl]methoxy-hydroxyphosphoryl]oxy-5-(5-methyl-2,4-dioxopyrimidin-1-yl)oxolan-2-yl]methoxy-hydroxyphosphoryl]oxy-5-(6-aminopurin-9-yl)oxolan-2-yl]methoxy-hydroxyphosphoryl]oxy-5-(6-aminopurin-9-yl)oxolan-2-yl]methoxy-hydroxyphosphoryl]oxy-5-(6-aminopurin-9-yl)oxolan-2-yl]methoxy-hydroxyphosphoryl]oxy-5-(6-aminopurin-9-yl)oxolan-2-yl]methoxy-hydroxyphosphoryl]oxyoxolan-2-yl]methoxy-hydroxyphosphoryl]oxy-5-(5-methyl-2,4-dioxopyrimidin-1-yl)oxolan-2-yl]methoxy-hydroxyphosphoryl]oxy-5-(4-amino-2-oxopyrimidin-1-yl)oxolan-2-yl]methoxy-hydroxyphosphoryl]oxy-5-(5-methyl-2,4-dioxopyrimidin-1-yl)oxolan-2-yl]methoxy-hydroxyphosphoryl]oxy-5-(5-methyl-2,4-dioxopyrimidin-1-yl)oxolan-2-yl]methoxy-hydroxyphosphoryl]oxy-5-(6-aminopurin-9-yl)oxolan-2-yl]methoxy-hydroxyphosphoryl]oxy-5-(6-aminopurin-9-yl)oxolan-2-yl]methoxy-hydroxyphosphoryl]oxy-5-(4-amino-2-oxopyrimidin-1-yl)oxolan-2-yl]methoxy-hydroxyphosphoryl]oxy-5-(4-amino-2-oxopyrimidin-1-yl)oxolan-2-yl]methoxy-hydroxyphosphoryl]oxy-5-(4-amino-2-oxopyrimidin-1-yl)oxolan-2-yl]methoxy-hydroxyphosphoryl]oxy-5-(6-aminopurin-9-yl)oxolan-2-yl]methoxy-hydroxyphosphoryl]oxy-5-(4-amino-2-oxopyrimidin-1-yl)oxolan-2-yl]methyl [5-(6-aminopurin-9-yl)-2-(hydroxymethyl)oxolan-3-yl] hydrogen phosphate Polymers Cc1cn(C2CC(OP(O)(=O)OCC3OC(CC3OP(O)(=O)OCC3OC(CC3O)n3cnc4c3nc(N)[nH]c4=O)n3cnc4c3nc(N)[nH]c4=O)C(COP(O)(=O)OC3CC(OC3COP(O)(=O)OC3CC(OC3COP(O)(=O)OC3CC(OC3COP(O)(=O)OC3CC(OC3COP(O)(=O)OC3CC(OC3COP(O)(=O)OC3CC(OC3COP(O)(=O)OC3CC(OC3COP(O)(=O)OC3CC(OC3COP(O)(=O)OC3CC(OC3COP(O)(=O)OC3CC(OC3COP(O)(=O)OC3CC(OC3COP(O)(=O)OC3CC(OC3COP(O)(=O)OC3CC(OC3COP(O)(=O)OC3CC(OC3COP(O)(=O)OC3CC(OC3COP(O)(=O)OC3CC(OC3COP(O)(=O)OC3CC(OC3CO)n3cnc4c(N)ncnc34)n3ccc(N)nc3=O)n3cnc4c(N)ncnc34)n3ccc(N)nc3=O)n3ccc(N)nc3=O)n3ccc(N)nc3=O)n3cnc4c(N)ncnc34)n3cnc4c(N)ncnc34)n3cc(C)c(=O)[nH]c3=O)n3cc(C)c(=O)[nH]c3=O)n3ccc(N)nc3=O)n3cc(C)c(=O)[nH]c3=O)n3cnc4c3nc(N)[nH]c4=O)n3cnc4c(N)ncnc34)n3cnc4c(N)ncnc34)n3cnc4c(N)ncnc34)n3cnc4c(N)ncnc34)O2)c(=O)[nH]c1=O JLCPHMBAVCMARE-UHFFFAOYSA-N 0.000 description 1
- 239000002671 adjuvant Substances 0.000 description 1
- 235000004279 alanine Nutrition 0.000 description 1
- 108010005233 alanylglutamic acid Proteins 0.000 description 1
- 108010087924 alanylproline Proteins 0.000 description 1
- 230000005875 antibody response Effects 0.000 description 1
- ODKSFYDXXFIFQN-UHFFFAOYSA-N arginine Natural products OC(=O)C(N)CCCNC(N)=N ODKSFYDXXFIFQN-UHFFFAOYSA-N 0.000 description 1
- 108010013835 arginine glutamate Proteins 0.000 description 1
- 108010068380 arginylarginine Proteins 0.000 description 1
- 108010077245 asparaginyl-proline Proteins 0.000 description 1
- 229940009098 aspartate Drugs 0.000 description 1
- 108010038633 aspartylglutamate Proteins 0.000 description 1
- 108010092854 aspartyllysine Proteins 0.000 description 1
- 108010068265 aspartyltyrosine Proteins 0.000 description 1
- 230000000903 blocking effect Effects 0.000 description 1
- 238000009835 boiling Methods 0.000 description 1
- 238000009395 breeding Methods 0.000 description 1
- 230000001488 breeding effect Effects 0.000 description 1
- 210000004899 c-terminal region Anatomy 0.000 description 1
- 238000005266 casting Methods 0.000 description 1
- 238000004113 cell culture Methods 0.000 description 1
- 229920002678 cellulose Polymers 0.000 description 1
- 239000001913 cellulose Substances 0.000 description 1
- 230000008859 change Effects 0.000 description 1
- 230000002860 competitive effect Effects 0.000 description 1
- 239000002299 complementary DNA Substances 0.000 description 1
- 238000007796 conventional method Methods 0.000 description 1
- NKLPQNGYXWVELD-UHFFFAOYSA-M coomassie brilliant blue Chemical compound [Na+].C1=CC(OCC)=CC=C1NC1=CC=C(C(=C2C=CC(C=C2)=[N+](CC)CC=2C=C(C=CC=2)S([O-])(=O)=O)C=2C=CC(=CC=2)N(CC)CC=2C=C(C=CC=2)S([O-])(=O)=O)C=C1 NKLPQNGYXWVELD-UHFFFAOYSA-M 0.000 description 1
- 235000018417 cysteine Nutrition 0.000 description 1
- XUJNEKJLAYXESH-UHFFFAOYSA-N cysteine Natural products SCC(N)C(O)=O XUJNEKJLAYXESH-UHFFFAOYSA-N 0.000 description 1
- 230000009089 cytolysis Effects 0.000 description 1
- 210000000805 cytoplasm Anatomy 0.000 description 1
- 238000013461 design Methods 0.000 description 1
- 238000012631 diagnostic technique Methods 0.000 description 1
- 201000010099 disease Diseases 0.000 description 1
- 208000037265 diseases, disorders, signs and symptoms Diseases 0.000 description 1
- 235000013399 edible fruits Nutrition 0.000 description 1
- 238000004520 electroporation Methods 0.000 description 1
- 210000001163 endosome Anatomy 0.000 description 1
- 108010078428 env Gene Products Proteins 0.000 description 1
- 108700004025 env Genes Proteins 0.000 description 1
- 238000001976 enzyme digestion Methods 0.000 description 1
- 229960003699 evans blue Drugs 0.000 description 1
- 230000003203 everyday effect Effects 0.000 description 1
- 238000002474 experimental method Methods 0.000 description 1
- 230000004927 fusion Effects 0.000 description 1
- 108700004026 gag Genes Proteins 0.000 description 1
- 108010027225 gag-pol Fusion Proteins Proteins 0.000 description 1
- 210000004392 genitalia Anatomy 0.000 description 1
- 239000003292 glue Substances 0.000 description 1
- 108010078144 glutaminyl-glycine Proteins 0.000 description 1
- VPZXBVLAVMBEQI-UHFFFAOYSA-N glycyl-DL-alpha-alanine Natural products OC(=O)C(C)NC(=O)CN VPZXBVLAVMBEQI-UHFFFAOYSA-N 0.000 description 1
- XBGGUPMXALFZOT-UHFFFAOYSA-N glycyl-L-tyrosine hemihydrate Natural products NCC(=O)NC(C(O)=O)CC1=CC=C(O)C=C1 XBGGUPMXALFZOT-UHFFFAOYSA-N 0.000 description 1
- 108010089804 glycyl-threonine Proteins 0.000 description 1
- 108010015792 glycyllysine Proteins 0.000 description 1
- 108010087823 glycyltyrosine Proteins 0.000 description 1
- 238000003306 harvesting Methods 0.000 description 1
- HNDVDQJCIGZPNO-UHFFFAOYSA-N histidine Natural products OC(=O)C(N)CC1=CN=CN1 HNDVDQJCIGZPNO-UHFFFAOYSA-N 0.000 description 1
- 108010025306 histidylleucine Proteins 0.000 description 1
- 108010085325 histidylproline Proteins 0.000 description 1
- 102000044890 human EPO Human genes 0.000 description 1
- 210000000987 immune system Anatomy 0.000 description 1
- 230000036039 immunity Effects 0.000 description 1
- 238000003018 immunoassay Methods 0.000 description 1
- 238000003125 immunofluorescent labeling Methods 0.000 description 1
- 229940031551 inactivated vaccine Drugs 0.000 description 1
- 230000005764 inhibitory process Effects 0.000 description 1
- AGPKZVBTJJNPAG-UHFFFAOYSA-N isoleucine Natural products CCC(C)C(N)C(O)=O AGPKZVBTJJNPAG-UHFFFAOYSA-N 0.000 description 1
- 229960000310 isoleucine Drugs 0.000 description 1
- 108010044374 isoleucyl-tyrosine Proteins 0.000 description 1
- 239000002523 lectin Substances 0.000 description 1
- 231100000518 lethal Toxicity 0.000 description 1
- 230000001665 lethal effect Effects 0.000 description 1
- 108010083708 leucyl-aspartyl-valine Proteins 0.000 description 1
- 239000007788 liquid Substances 0.000 description 1
- 229940023832 live vector-vaccine Drugs 0.000 description 1
- 244000144972 livestock Species 0.000 description 1
- 230000002934 lysing effect Effects 0.000 description 1
- 108010064235 lysylglycine Proteins 0.000 description 1
- 210000001161 mammalian embryo Anatomy 0.000 description 1
- 239000003550 marker Substances 0.000 description 1
- 230000007246 mechanism Effects 0.000 description 1
- 230000001404 mediated effect Effects 0.000 description 1
- 239000002609 medium Substances 0.000 description 1
- 229930182817 methionine Natural products 0.000 description 1
- 238000002941 microtiter virus yield reduction assay Methods 0.000 description 1
- 238000010369 molecular cloning Methods 0.000 description 1
- 108700004028 nef Genes Proteins 0.000 description 1
- 239000013642 negative control Substances 0.000 description 1
- 229940023146 nucleic acid vaccine Drugs 0.000 description 1
- 108020004707 nucleic acids Proteins 0.000 description 1
- 102000039446 nucleic acids Human genes 0.000 description 1
- 150000007523 nucleic acids Chemical class 0.000 description 1
- 238000002515 oligonucleotide synthesis Methods 0.000 description 1
- 244000052769 pathogen Species 0.000 description 1
- 229940049954 penicillin Drugs 0.000 description 1
- COLNVLDHVKWLRT-UHFFFAOYSA-N phenylalanine Natural products OC(=O)C(N)CC1=CC=CC=C1 COLNVLDHVKWLRT-UHFFFAOYSA-N 0.000 description 1
- 239000013600 plasmid vector Substances 0.000 description 1
- 229910052697 platinum Inorganic materials 0.000 description 1
- 108700004029 pol Genes Proteins 0.000 description 1
- 238000006116 polymerization reaction Methods 0.000 description 1
- 239000013641 positive control Substances 0.000 description 1
- 239000000843 powder Substances 0.000 description 1
- 108010070643 prolylglutamic acid Proteins 0.000 description 1
- 108010029020 prolylglycine Proteins 0.000 description 1
- 239000002994 raw material Substances 0.000 description 1
- 230000004044 response Effects 0.000 description 1
- 238000000926 separation method Methods 0.000 description 1
- 108010026333 seryl-proline Proteins 0.000 description 1
- 108010071207 serylmethionine Proteins 0.000 description 1
- 241000894007 species Species 0.000 description 1
- 239000012192 staining solution Substances 0.000 description 1
- 239000011550 stock solution Substances 0.000 description 1
- 229960005322 streptomycin Drugs 0.000 description 1
- 239000013589 supplement Substances 0.000 description 1
- 230000004083 survival effect Effects 0.000 description 1
- 208000011580 syndromic disease Diseases 0.000 description 1
- 108010061238 threonyl-glycine Proteins 0.000 description 1
- 238000013518 transcription Methods 0.000 description 1
- 230000035897 transcription Effects 0.000 description 1
- 230000009466 transformation Effects 0.000 description 1
- 230000010474 transient expression Effects 0.000 description 1
- 230000014621 translational initiation Effects 0.000 description 1
- 230000005945 translocation Effects 0.000 description 1
- 108010038745 tryptophylglycine Proteins 0.000 description 1
- 108010044292 tryptophyltyrosine Proteins 0.000 description 1
- OUYCCCASQSFEME-UHFFFAOYSA-N tyrosine Natural products OC(=O)C(N)CC1=CC=C(O)C=C1 OUYCCCASQSFEME-UHFFFAOYSA-N 0.000 description 1
- 108010051110 tyrosyl-lysine Proteins 0.000 description 1
- 241001430294 unidentified retrovirus Species 0.000 description 1
- 239000012646 vaccine adjuvant Substances 0.000 description 1
- 229940124931 vaccine adjuvant Drugs 0.000 description 1
- 238000012795 verification Methods 0.000 description 1
Images
Landscapes
- Medicines Containing Antibodies Or Antigens For Use As Internal Diagnostic Agents (AREA)
Abstract
本发明涉及一种人工合成的含有鸡体偏嗜性密码子的基因optiHA，其阅读框架含1707bp核苷酸，共编码568个氨基酸，该基因与H5亚型高致病力禽流感病毒A/Goose/GuangDong/1/96(H5N1)[GD/1/96(H5N1)]血凝素(HA)基因的核苷酸同源率为70％，氨基酸同源率为100％，编码H5亚型禽流感病毒GD/1/96(H5N1)的血凝素(HA)蛋白。本发明同时涉及该基因作为H5亚型流感DNA疫苗及其它基因工程疫苗的免疫原基因的应用。
Description
技术领域
本发明涉及一种人工合成的含有鸡体偏嗜性密码子的基因optiHA，该基因与H5亚型高致病力禽流感病毒A/Goose/GuangDong/1/96(H5N1)[GD/1/96(H5N1)]血凝素(HA)基因的核苷酸同源率为70％，氨基酸同源率为100％，编码并表达H5亚型禽流感病毒GD/1/96(H5N1)的血凝素(HA)蛋白。该基因可作为H5亚型流感DNA疫苗及其它基因工程疫苗的免疫原基因。
背景技术
禽流感(Avian Influenza，AI)是由禽流感病毒(Avian Influenza Virus，AIV)引起的禽类感染和/或疾病综合征，AIV在分类学上属于：病毒界(Vira)----正粘病毒科(Orthomyxoviridae)----流感病毒属(Influenza VirusA and B)----禽流感病毒(Avian Influenza Virus)。
高致病力禽流感(Highly Pathogenic Avian Influenza，HPAI)被国际兽医局列为A类烈性传染病(Alexander，D.J，et al.An outbreak of highlypathogenic avian influenza in turkeys in Great Britain in 1991.VeterinaryRecord，1993，132，535-536.)。近年来发现，H5和H7亚型高致病力禽流感病毒(Highly Pathogenic Avian Influenza Virus，HPAIV)不但可给养禽业造成巨大损失，而且可以感染人并致人死亡，对HPAI的防制是关系养禽业健康发展与人类生命安全的重大课题(Tran TH，et al.Avian influenza A(H5N1)in 10 patients in Vietnam.N Engl J Med.2004，350(12)：1179-88.)。DNA疫苗是防制高致病力禽流感最具潜力的基因工程疫苗之一，其抗原合成和提呈过程与病原的自然感染相似，抗原蛋白经内源性合成后转运到细胞表面，通过MHC-I类MHC-II类分子直接递呈给免疫系统，能够诱导机体产生特异性CTL反应(Robinson HL.Nucleic acid vaccines：an overview.Vaccine，1997，15：785-787)。
血凝素(HA)是禽流感病毒主要的免疫原蛋白，它可诱导机体产生抗体介导的特异性体液免疫应答，抗HA的抗体可以通过干扰病毒与唾液酸受体的结合或者病毒囊膜与内吞体膜的融合过程从而中和病毒的感染。大量研究证明基于流感病毒HA基因的DNA疫苗可以有效保护同一HA亚型流感病毒的攻击(Robinson HL，et al.Protection against a lethal influenzavirus challenge by immunization with a haemagglutinin-expressing plasmidDNA.Vaccine 1993，11(9)：957-60.2.Kodihalli S，et al.DNA vaccine encodinghemagglutinin provides protective immunity against H5N1 influenza virusinfection in mice.Virol.1999 Mar，73(3)：2094-8.3.Kodihalli S，et al.Strategies for inducing protection against avian influenza A virus subtypes withDNA vaccines.Vaccine.2000 May 22，18(23)：2592-9.3)。A/Goose/GuangDong/1/96(H5N1)[GD/1/96(H5N1)]为我国最早分离的H5亚型高致病力禽流感病毒(唐秀英等.中国禽流感流行株的鉴定.中国畜禽传染病，1998，20(1)：1-5.)。研究表明，表达GD/1/96(H5N1)HA基因的DNA疫苗质粒pCIHA不仅表现出了良好的免疫原性，而且诱导产生的有效保护性HI抗体可以持续至一年(H Chen，et al.DNA immunization elicitshigh HI antibody and protects chicken from AIV challenge.ImmuntionalCongress Series 1219(2001)917-921.姜永萍.抗原基因密码子及其表达载体的优化增强H5亚型禽流感DNA疫苗的免疫保护效果[博士学位论文].哈尔滨，中国农业科学院哈尔滨兽医研究所，2004.)，但由于H5亚型禽流感DNA疫苗高剂量的依赖性造成使用成本提高，与传统禽流感灭活疫苗相比经济可行性缺乏竞争力。将异源的免疫原基因的密码子优化为免疫接种动物偏嗜使用的密码子可显著提高DNA疫苗抗原表达效率、降低有效免疫剂量(Garmory HS，et al.DNA vaccines：improving expression ofantigens.Genet Vaccines Ther.2003 Sep 16，1(1)：2.)，并将有可能突破DNA疫苗进一步推广应用的制约瓶颈。
发明内容
本发明的一个目的是提供一种人工合成的密码子优化的基因optiHA(SEQ ID NO：1)，该基因含有鸡体偏嗜使用的密码子，编码并表达H5亚型禽流感病毒A/Goose/GuangDong/1/96(H5N1)[GD/1/96(H5N1)]的血凝素(HA)蛋白(SEQ ID NO：2)，该基因可作为H5亚型禽流感DNA疫苗及其它基因工程疫苗的免疫原基因。
本发明的另一个目的是提供一种包含optiHA基因的表达载体。
本发明的另一个目的是提供一种用于表达optiHA基因的宿主细胞。所述的宿主细胞可以是人胚胎肾细胞、COS细胞、鸡胚成纤维细胞等真核细胞。
本发明还涉及optiHA基因在制备H5亚型流感病毒DNA疫苗中作为免疫原基因的应用，在亚单位疫苗、重组禽痘病毒基因工程疫苗等基因工程疫苗中作为免疫原基因的应用，以及在单克隆抗体制备中作为免疫原基因的应用。
本发明关于H5亚型禽流感DNA疫苗免疫原基因的密码子优化策略的理论依据有以下几点：
1)禽流感病毒表面结构蛋白HA是其最主要的免疫原蛋白。
2)鸡与A型流感病毒保护性免疫原HA蛋白氨基酸密码子使用频率存在着明显差异。编码氨基酸密码子的摇摆性导致了不同物种间细胞在蛋白翻译过程中具有密码子使用的偏嗜性。由于削弱的翻译效率，特别是相应的tRNA含量的缺少和起始密码子的改变，将会大大影响蛋白质的翻译起始和延伸效率(Hamdan FF，et al.Codon optimization improves heterologousexpression of a Schistosoma mansoni cDNA in HEK293 cells.Parasitol Res.2002 Jun，88(6)：583-6.Epub 2002 Feb 16.)。鸡与哺乳动物细胞在蛋白翻译过程中具有相同的密码子偏嗜性，但与A型流感病毒保护性免疫原HA蛋白氨基酸密码子使用频率存在着明显差异，在20个氨基酸中，只有6个氨基酸使用了相同的密码子，其余14个氨基酸的密码子使用情况截然不同(Holm L.Codon usage and gene expression.Nucleic Acids Res.1986 Apr11，14(7)：3075-87.)。
3)将异源的免疫原基因的密码子优化为免疫接种动物偏嗜使用的密码子可显著提高抗原表达效率。将异源的免疫原基因的密码子优化为免疫接种动物偏嗜使用的密码子可显著提高抗原表达效率，这一结果已在人类免疫缺陷病(艾滋病)的DNA疫苗研究中已得到了充分的证实。鉴于大多数慢病毒结构蛋白编码基因密码子使用频率严重偏离哺乳动物细胞密码子的偏嗜性，Haas J(Haas J，et al.Codon usage limitation in theexpression of HIV-1 envelope glycoprotein.Curr Biol.1996 Mar1，6(3)：315-24.)等利用合成的末端重叠寡核苷酸，采用重叠PCR技术扩增了哺乳动物密码子优化的人类免疫缺陷病毒(HIV)MN株gp120基因，大大提高了体外表达水平，用于构建DNA疫苗，显著增强了特异性的细胞和体液免疫反应水平。除Env糖蛋白，密码子优化的HIVGag、Gag-pol、Tat、Rev以及Nef用于DNA疫苗或重组病毒活载体疫苗同样取得了显著改善的特异细胞和体液免疫反应(Gao F，et al.Codon usage optimization ofHIV type 1 subtype C gag，pol，env，and nef genes：in vitro expression andimmune responses in DNA-vaccinated mice.AIDS Res Hum Retroviruses.2003 Sep，19(9)：817-23.)。目前几乎所有的HIV重组疫苗都采用了密码子优化的免疫原基因。密码子优化的免疫原基因在人的乳头瘤病毒(Cid-Arregui A，et al.A synthetic E7 gene of human papillomavirus type 16that yields enhanced expression of the protein in mammalian cells and is usefulfor DNA immunization studies.J Virol.2003 Apr；77(8)：4928-37)、狗的乳头瘤病毒(Moore RA，et al.Intraepithelial DNA immunisation with a plasmidencoding a codon optimised COPV E1 gene sequence，but not the wild-typegene sequence completely protects against mucosal challenge with infectiousCOPV in beagles.Virology.2002 Dec 20；304(2)：451-9.)以及SARS冠状病毒(Gao W，et al.Effects of a SARS-associated coronavirus vaccine in monkeys.Lancet.2003 Dec 6；362(9399)：1895-6.)DNA疫苗中相继得到应用。密码子优化的免疫原基因增强免疫反应的机制不仅仅涉及翻译过程密码子移动速度加快。密码子哺乳动物偏嗜性优化往往导致mRNA中CG含量的大幅度提高，因而彻底改变RNA的二级结构。这一方面有可能增加mRNA的稳定性，另一方面还有可能增加mRNA转录后从细胞核向细胞浆的转运效率。另外，CG含量的增加还有可能增加CpG的出现频率，进一步发挥DNA疫苗本身免疫佐剂的作用(McCluskie MJ，et al.The use of CpG DNAas a mucosal vaccine adjuvant.Curr Opin Investig Drugs.2001Jan；2(1)：35-9.)。
4)重叠延伸PCR[Overlap Extension PCR(OE-PCR)]技术是免疫原基因密码子优化的最简单、基本的策略。重叠延伸PCR技术的基本原理是将相互互补重叠的两个片段(一般基因合成公司可合成长为100bp左右的寡核苷酸片段)置于同一PCR反应体系中，控制PCR条件使其互补延伸后再加入片段两边的引物(第一个片段的上游引物与第二个片段的下游引物)继续进行PCR反应而得到含有两个片段的大片段(Mehta RK，et al.Bridge-overlap-extension PCR method for constructing chimeric genes.Biotechniques.1999 Jun，26(6)：1082-6.)。对于免疫原基因的密码子优化，Haas J(Haas J，et al.Codon usage limitation in the expression of HIV-1envelope glycoprotein.Curr Biol.1996 Mar 1，6(3)：315-24.)等利用重叠的引物及特异的酶切位点，采用PCR技术扩增了gp120的8个长的寡核苷酸片段，然后将这8个片段以一定的顺序亚克隆到载体pCdm7上而合成和构建了gp120基因，此后这一技术被大多数研究者所采用。Kim CH(KimCH，et al.Codon optimization for high-level expression of humanerythropoietin(EPO)in mammalian cells.Gene.1997 Oct15，199(1-2)：293-301.)等利用重叠延伸PCR技术(OE-PCR)技术合成了人成熟的促红细胞生成素基因(EPO)。合成的8个长为80-90bp的寡核苷酸片段互相互补重叠15-20bp以便于PCR反应。首先将合成的前4个寡核苷酸片段及后4个寡核苷酸片段分别置于同一PCR反应体系中进行PCR，通过第一轮PCR，分别扩增得到了编码EPO基因的N末端半个片段和C末端半个片段，然后将两个PCR反应体系得到的小片段混合，并以与前面同样的PCR反应条件进行5个循环，然后将含有酶切位点和EPO基因的5‘和3’端互补接头序列的引物加入，再进一步扩增30个循环而得到了全长的EPO基因。
基于以上几点，我们以寡核苷酸合成、重叠延伸PCR(OverlapExtension PCR，OE-PCR)技术、限制性内切酶酶切和连接技术以及测序技术获得一种新的密码子优化的基因optiHA，该基因与A/Goose/GuangDong/1/96(H5N1)[GD/1/96(H5N1)]HA基因的核苷酸同源率为70％，氨基酸同源率为100％，编码H5亚型禽流感病毒GD/1/96(H5N1)的HA蛋白。将该基因和禽流感病毒A/Goose/GuangDong/1/96(H5N1)[GD/1/96(H5N1)]的HA基因分别插入CMV启动子表达载体pCI，构建的H5亚型禽流感DNA疫苗质粒pCIoptiHA(含基因optiHA)和pCIHA(含GD/1/96(H5N1)HA基因)分别转染293T细胞，间接免疫荧光和Western-blot检测证实密码子优化的HA基因optiHA的HA蛋白体外瞬时表达水平显著高于野生型HA基因。随之将pCIoptiHA及pCIHA分别以100μg和10μg剂量一次免疫3周龄SPF鸡，4周后用100LD50的HPAIV GD/1/96(H5N1)鼻腔途径进行攻击，结果免疫SPF鸡后，100μg pCIoptiHA可形成75％的免疫保护，100μgpCIHA可形成50％的免疫保护；10μg pCIoptiHA可形成对免疫鸡75％的部分免疫保护，而10μg pCIHA则基本不能形成免疫保护；pCIoptiHA诱导的HI抗体水平远远高于pCIHA。结果表明，HA基因密码子的优化可显著提高HA基因体外表达水平和H5亚型禽流感DNA疫苗诱导的保护性抗体免疫反应水平，提高免疫原性，增强免疫保护效果。
附图说明
图1为按照本发明的optiHA基因的核苷酸序列及对应的氨基酸序列。
图2为片段optiHAF1-optiHAF11的PCR扩增产物电泳图片，其中泳道1为DNA MarkerDL2000(D501A，宝生物技术工程(大连)有限公司)；泳道2为片段optiHAF1的PCR产物，大小为88bp；泳道3为片段optiHAF2的PCR产物，大小为162bp；泳道4为片段optiHAF3的PCR产物，大小为204bp；泳道5为片段optiHAF4的PCR产物，大小为162bp；泳道6为片段optiHAF5的PCR产物，大小为164bp；泳道7为片段optiHAF6的PCR产物，大小为201bp；泳道8为片段optiHAF7的PCR产物，大小为201bp；泳道9为片段optiHAF8的PCR产物，大小为155bp；泳道10为片段optiHAF9的PCR产物，大小为157bp；泳道11为片段optiHAF10的PCR产物，大小为168bp；泳道12为片段optiHAF11的PCR产物，大小为182bp。
图3为基因optiHA的连接构建策略，optiHA基因的11个片段optiHAF1-optiHAF11分别由合成的寡核苷酸片段利用重叠延伸PCR法扩增并克隆到pBluscript载体上，之后将11个片段的克隆质粒pBluscript-optiHAF1-pBluscriptoptiHAF11利用选定的酶切位点以一定的顺序相连而构建出optiHA基因。
图4为基因optiHA的PCR鉴定图片，其中泳道1为DNA MarkerDL2000；泳道2为PCR鉴定时的H2O对照，泳道3为基因optiHA的1.8kb大小的PCR产物。
图5为基因optiHA的酶切鉴定图片，其中泳道1为DNA 1KbMarker(CD002，北京驼峰星生物技术工程有限公司)；泳道2为含有基因optiHA的质粒pCIoptiHAFull用PvuII酶切时得到的1.0Kb和4.8kb的两条带；泳道3为质粒pCIoptiHAFull用XhoI酶切时得到的1.6kb和4.2kb的条带；泳道4为质粒pCIoptiHAFull用NotI酶切时得到的1.2kb和4.6kb的条带；泳道5为质粒pCIoptiHAFull用EcoRI酶切时得到的线性化的5.8kb的片段；泳道6为质粒pCIoptiHAFull用BamHI酶切时得到的300bp和5.5kb的条带；泳道7为DNA Marker DL15000。
图6为质粒pCIoptiHA的酶切分析图谱，其中泳道1为DNA 1KbMarker(CD002，北京驼峰星生物技术工程有限公司)；泳道2为pCIoptiHA用EcoRI酶切时得到的线性化的5.8kb的片段；泳道3为pCIoptiHA用PstI酶切时得到的1.2Kb、530bp、300bp和3.87kb的条带、泳道4为pCIoptiHA用BglII酶切得到的1.1kb和4.7kb的条带。
图7为表达A/Goose/GuangDong/1/96(H5N1)[GD/1/96(H5N1)]的HA基因的质粒pCIHA(作为对照)转染293T细胞后24h在荧光显微镜(200×)下观察到的间接免疫荧光检测结果，表达HA蛋白的细胞主要在其细胞膜呈现为亮绿色，图中箭头所示的细胞为表达HA蛋白的细胞。
图8为表达optiHA基因的质粒pCIoptiHA转染293T细胞后24h在荧光显微镜(200×)下观察到的间接免疫荧光检测结果，表达HA蛋白的细胞主要在其细胞膜呈现为亮绿色，图中箭头所示的细胞为表达HA蛋白的细胞。
图9为表达A/Goose/GuangDong/1/96(H5N1)[GD/1/96(H5N1)]的HA基因的质粒pCIHA(作为对照)转染293T细胞后48h在荧光显微镜(200×)下观察到的间接免疫荧光检测结果，表达HA蛋白的细胞主要在其细胞膜呈现为亮绿色，图中箭头所示的细胞为表达HA蛋白的细胞。
图10为表达optiHA基因的质粒pCIoptiHA转染293T细胞后48h在荧光显微镜(200×)下观察到的间接免疫荧光检测结果，表达HA蛋白的细胞主要在其细胞膜呈现为亮绿色，图中箭头所示的细胞为表达HA蛋白的细胞。
图11为293T细胞对照在荧光显微镜(200×)下观察到的间接免疫荧光检测结果，所有细胞均没有表达HA蛋白，细胞在荧光显微镜下不呈现亮绿色。
图12为pCIoptiHA和pCIHA的SDS-PAGE图片，其中泳道1为转染质粒pCIHA的293T细胞；泳道2为转染质粒pCIoptiHA的293T细胞；泳道3为H5亚型禽流感病毒A/Goose/GuangDong/1/96(H5N1)[GD/1/96(H5N1)]；泳道4为293T细胞对照；泳道5为蛋白分子量标准。
图13为pCIoptiHA和pCIHA的Western blot分析结果图片，其中泳道1为293T细胞对照；泳道2为H5亚型禽流感病毒A/Goose/GuangDong/1/96(H5N1)[GD/1/96(H5N1)]的大小为74KDa的HA蛋白；泳道3为质粒pCIoptiHA在293T细胞中表达的大小为74KDa的HA蛋白；泳道4为质粒pCIHA在293T细胞中表达的大小为74KDa的HA蛋白。
具体实施方式
下文将参考实施例详细描述本发明，所述实施例仅是意图举例说明本发明，而不是意图限制本发明的范围。本发明的范围由后附的权利要求具体限定。
本发明实施例中所用原料及试验试剂的来源：
(1)H5亚型高致病力禽流感病毒(HPAIV)A/Goose/Guangdong/1/96(H5N1)[简写为GD/1/96(H5N1)]购自哈尔滨兽医研究所。
(2)A/Goose/Guangdong/1/96(H5N1)[GD/1/96(H5N1)]的HA基因核苷酸序列来自Genebank(Genebank登录号：AF144305)，其氨基酸序列由核苷酸序列推导而来。
(3)293T细胞为人胚胎肾细胞(CRL-11268)，来自ATCC。
(4)受体菌E.coli JM109(D9025)为宝生物技术工程(大连)有限公司产品。
(5)质粒pBluscript(212207)为Stratagene公司产品。
(6)表达载体pCI(E1731，Genebank登录号：U47119)为Promega公司产品。
(7)PlatinumPfx Taq DNA聚合酶(11708-013)为Ivitrogen公司产品。
(8)T4DNA聚合酶(M0203S)、T4DNA连接酶(M0202S)为New EnglandBliolabs公司产品。
(9)SapI(R0569S)为New England Bliolabs公司产品，BglII(D1021A)、PstI(D1073A)、PvuII(D1076A)、EcoO109I(D1043A)、NotI(D1166A)、SphI(D1180A)、KpnI(D1068A)、XhoI(D1094A)、EcoRI(D1040A)和XbaI(D1093A)等限制性内切酶均为宝生物技术工程(大连)有限公司产品。
(10)胶回收(小量)试剂盒(W5212)、质粒(小量)抽提试剂盒(W5002)为上海华舜生物工程有限公司产品。
(11)CEQTM DTCS-QUICK Star Kit(608120)为Beckman公司产品。
(12)PerfectprepPlasmid Mini(0032005.500)为EppendorfAG公司产品。
(13)LipofectamineTM2000(11668-027)为Invitrogen公司产品。
(14)DMEM干粉(12100-046)为GIBCO公司产品，用无离子水配制成pH7.2的1×DMEM(配制1000ml培养液需加入3.7g的NaHCO3和2.4g的HEPES)溶液4℃保存，临用时加入终浓度为10％的胎牛血清(26149-095，Gibco)、100U/ml青霉素和100μg/ml链霉素。
(15)OPTi-MEMI(51985-034)为不含血清的用于转染的培养液，为Invitrogen公司产品。
(16)H5亚型禽流感多克隆阳性血清(批号：040524)为哈尔滨兽医研究所产品。
(17)蛋白电泳分子量标准(SM0431)为晶美生物工程有限公司产品。
(18)荧光标记的二抗兔抗鸡IgG(F8888，FITC-IgG)为Sigma公司产品。
(19)辣根过氧化物酶标记的二抗兔抗鸡IgG(A9046，HRPO-IgG)为Sigma公司产品。
(20)DAB显色试剂盒(AR1022)为武汉博士德生物工程有限公司产品。
(21)9～11日龄鸡胚购自哈尔滨兽医研究所试验动物中心。
(22)无特定病原菌(Specific-pathogen-free，SPF)的白色来杭鸡购自哈尔滨兽医研究所试验动物中心。
(23)其它化学试剂均为上海华舜生物工程有限公司产品。
实施例1.H5亚型HA基因鸡体偏嗜性密码子的改写与序列分析
利用密码子使用频率表(见附表1，国家科学数字图书馆，生命科学学科信息门户，密码子使用数据库，网址为http：//www.kazusa.or.jp/codon/)，将H5亚型禽流感病毒A/Goose/Guangdong/1/96(H5N1)[GD/1/96(H5N1)]HA基因的密码子全部变换为鸡体偏嗜性密码子，并书写出密码子优化后的HA基因(命名为optiHA)的序列，长1779bp(SEQ ID NO：1，参见图1)，并对optiHA基因序列用DNASTAR软件(DNASTAR.Inc.)进行序列和酶切位点分析，根据optiHA基因上可以选择利用的单酶切位点，将optiHA基因分成11个长约200bp的片段，11个片段依次命名为optiHAF1(片段长88bp，在基因optiHA上的位置为1-88)、optiHAF2(片段长162bp，在基因optiHA上的位置为81-243)、optiHAF3(片段长204bp，在基因optiHA上的位置为237-441)、optiHAF4(片段长162bp，在基因optiHA上的位置为436-597)、optiHAF5(片段长164bp，在基因optiHA上的位置为587-751)、optiHAF6(片段长201bp，在基因optiHA上的位置为744-944)、optiHAF7(片段长201bp，在基因optiHA上的位置为939-1139)、optiHAF8(片段长155bp，在基因optiHA上的位置为1134-1288)、optiHAF9(片段长157bp，在基因optiHA上的位置为1285-1441)、optiHAF10(片段长168bp，在基因optiHA上的位置为1436-1603)、optiHAF11(片段长182bp，在基因optiHA上的位置为1598-1779)。这11个片段在选定的单酶切位点相互重叠并各自包含选定的单酶切位点，即片段optiHAF1的3‘端和片段optiHAF2的5’端相互重叠含有BglII位点；片段optiHAF2的3‘端和片段optiHAF3的5’端相互重叠含有PstI位点；片段optiHAF3的3‘端和片段optiHAF4的5’端相互重叠含有PvuII位点：片段optiHAF4的3‘端和片段optiHAF5的5’端相互重叠含有EcoO109I位点；片段optiHAF5的3‘端和片段optiHAF6的5’端相互重叠含有NotI位点；片段optiHAF6的3‘端和片段optiHAF7的5’端相互重叠含有SphI位点；片段optiHAF7的3‘端和片段optiHAF8的5’端相互重叠含有KpnI位点；片段optiHAF8的3‘端和片段optiHAF9的5’端由于没有可以选择的的单酶切位点而分别在片段optiHAF8的3‘端和片段optiHAF9的5’端另外加入了SapI位点识别序列(该加入的酶切位点不包含在图1的序列中)；片段optiHAF9的3‘端和片段optiHAF10的5’端相互重叠含有PvuII位点；片段optiHAF10的3‘端和片段optiHAF11的5’端相互重叠含有XhoI。11个片段中，片段optiHAF1设计直接由一段正向(5’-3’)的寡核苷酸片段利用OE-PCR法扩增合成，片段optiHAF2-optiHAF11分别由两个正反两向、互相重叠约20bp、长约100bp的寡核苷酸片段利用OE-PCR法扩增合成。
附表1 鸡和流感病毒最偏嗜的密码子
氨基酸 鸡 流感病毒
精氨酸Arg(R) CGC AGA
亮氨酸Leu(L) CUG CUG
丝氨酸Ser(S) AGC UCA
苏氨酸Thr(T) ACC ACA
脯氨酸Pro(P) CCC CCA
丙氨酸Ala(A) GCC GCA
甘氨酸Gly(G) GGC GGA
缬氨酸Val(V) GUG GUG
赖氨酸Lys(K) AAG AAA
谷氨酰胺Gln(Q) CAG CAA
天冬酰胺Asn(N) AAC AAU
组氨酸His(H) CAC CAU
谷氨酸Glu(E) GAG GAA
天冬氨酸Asp(D) GAC GAU
酪氨酸Tyr(Y) UAC UAU
半胱氨酸Cys(C) UGC UGC
苯丙氨酸Phe(F) UUC UUC
异亮氨酸Ile(I) AUC AUA
甲硫氨酸Met(M) AUG AUG
色氨酸Trp(W) UGG UGG
实施例2.用于OE-PCR的寡核苷酸片段和引物的合成
片段optiHAF1的寡核苷酸片段optiHAF1-Forward(101)直接送由Sigma公司合成；片段optiHAF2的两个寡核苷酸片段为optiHAF2-Forward(90)和optiHAF2-Reverse(101)；片段optiHAF3的两个寡核苷酸片段为optiHAF3-Forward(110)和optiHAF3-Reverse(110)；片段optiHAF4的两个寡核苷酸片段为optiHAF4-Forward(99)和optiHAF4-Reverse(100)；片段optiHAF5的两个寡核苷酸片段为optiHAF5-Forward(97)和optiHAF5-Reverse(99)；片段optiHAF6的两个寡核苷酸片段为optiHAF6-Forward(110)和optiHAF6-Reverse(110)；片段optiHAF7的两个寡核苷酸片段为optiHAF7-Forward(110)和optiHAF7-Reverse(110)；片段optiHAF8的两个寡核苷酸片段为optiHAF8-Forward(91)和optiHAF8-Reverse(103)；片段optiHAF9的两个寡核苷酸片段为optiHAF9-Forward(97)和optiHAF9-Reverse(99)；片段optiHAF10的两个寡核苷酸片段为optiHAF10-Forward(99)和optiHAF10-Reverse(101)；片段optiHAF11的两个寡核苷酸片段为optiHAF11-Forward(107)和optiHAF11-Reverse(101)。这些寡核苷酸片段均送由Sigma公司合成(见附表2)。
附表2 合成的寡核苷酸片段
片段编号 寡核苷酸片段序列
optiHAF1- AGCGAAAGCAGGGGTCCAATCTGTCAAAATGGAGA
Forward(101) AGATCGTGCTGCTGCTGGCCATCGTGAGCCTGGTGA
AGAGCGACCAGATCTGCAT CGGCTACCACG (SEQ
ID NO：3)
CCAGATCTGCATCGGCTACCACGCCAACAACAGCAC
optiHAF2-
CGAGCAGGTGGACACCATCATGGAGAAGAACGTGAC
Forward(90)
CGTGACCCACGCCCAGGA (SEQ ID NO：4)
GCCACGCTGCAGTCGCGCAGGATCAGGGGCTTCACG
optiHAF2- CCGTTCAGGTCGCACAGCTTGCCGTTGTGGGTCTTCT
Reverse(101) CCAGGATGTCCTGGGCGTGGGTCACGGT (SEQ ID
NO：5)
TGCAGCGTGGCCGGCTGGCTGCTGGGCAACCCCATG
optiHAF3- TGCGACGAGTTCATCAACGTGCCCGAGTGGAGCTAC
Forward(110) ATCGTGGAGAAGGCCAGCCCCGCCAACGACCTGTGC
TA (SEQ ID NO：6)
CTGCTCTTGGGGATGATCTGGATCTTCTCGAAGTGGT
optiHAF3- TGGTGCGGCTCAGCAGGTGCTTCAGCTCCTCGTAGTC
Reverse(110) GTTGAAGTCGCCGGGGTAGCACAGGTCGTTGGCGGG
(SEQ ID NO：7)
CAAGAGCAGCTGGAGCAACCACGACGCCAGCAGC
optiHAF4- GGCGTGAGCAGCGCCTGCCCCTACCACGGCCGCAGC
Forward(99) AGCTTCTTCCGCAACGTGGTGTGGCTGA (SEQ ID
NO：8)
TGCCCCACAGCACCAGCAGGTCCTCCTGGTTGGTGT
optiHAF4- TGTTGTAGCTGCGCTTGATGGTGGGGTAGGCGCTGTT
Reverse(100) CTTCTTGATCAGCCACACCACGTTGCG (SEQ ID
NO：9)
AGGAGGACCTGCTGGTGCTGTGGGGCATCCACCACC
optiHAF5-
CCAACGACGCCGCCGAGCAGACCAAGCTGTACCAGA
Forward(97)
ACCCCACCACCTACATCAGCGTGGG (SEQ ID NO：10)
optiHAF5- CTCCATGCGGCCGCTCTGCCCGTTCACCTTGGGGCGG
Reverse(99) GTGGCGATCTCGGGCACCAGGCGCTGGTTCAGGGTG
CTGGTGCCCACGCTGATGTAGGTGGT(SEQ ID NO：11)
GCGGCCGCATGGAGTTCTTCTGGACCATCCTGAAGC
optiHAF6- CCAACGACGCCATCAACTTCGAGAGCAACGGCAACT
Forward(110) TCATCGCCCCCGAGTACGCCTACAAGATCGTGAAGA
AG (SEQ ID NO：12)
CATGCTGCTGTTGATGGCGCCCATGGGGGTCTGGCAC
optiHAF6- TTGGTGTTGCAGTTGCCGTACTCCAGCTCGCTCTTCA
Reverse(110) TGATGGCGCTGTCGCCCTTCTTCACGATCTTGTAGG
(SEQ ID NO：13)
GCATGCCCTTCCACAACATCCACCCCCTGACCATCGG
optiHAF7- CGAGTGCCCCAAGTACGTGAAGAGCAACCGCCTGGT
Forward(110) GCTGGCCACCGGCCTGCGCAACACCCCCCAGCGCGA
G (SEQ ID NO：14)
GTACCCGTACCAGCCGTCCACCATGCCCTGCCAGCCG
optiHAF7- CCCTCGATGAAGCCGGCGATGGCGCCGAACAGGCCG
Reverse(110) CGCTTCTTGCGGCGGCGCTCGCGCTGGGGGGTGTTG
C (SEQ ID NO：15)
GGTACGGGTACCACCACAGCAACGAGCAGGGCAGC
optiHAF8-
GGCTACGCCGCCGACAAGGAGAGCACCCAGAAGGC
Forward(91)
CATCGACGGCGTGACCAACAA (SEQ ID NO：16)
GCGGCGGCTCTTCCTCCAGGTTGTTGAACTCGCGGC
optiHAF8- CCACGGCCTCGAACTGGGTGTTCATCTTGTCGATGAT
Reverse(103) GCTGTTCACCTTGTTGGTCACGCCGTCGAT (SEQ ID
NO：17)
CAACCTGCTCTTCGGAGCGCCGCATCGAGAACCTGA
optiHAF9-
ACAAGCAGATGGAGGACGGCTTCCTGGACGTGTGGA
Forward(97)
CCTACAACGCCGAGCTGCTGGTGCT(SEQ ID NO：18)
GCGCAGCTGCAGGCGCACCTTGTCGTACAGGTTCTT
optiHAF9-
CACGTTGCTGTCGTGGAAGTCCAGGGTGCGCTCGTT
Reverse(99)
CTCCATCAGCACCAGCAGCTCGGCGTT (SEQ
ID NO：19)
GTGCGCCTGCAGCTGCGCGACAACGCCAAGGAGCTG
optiHAF10- GGCAACGGCTGCTTCGAGTTCTACCACAAGTGCGAC
Forward(99) AACGAGTGCATGGAGAGCGTGAAGAAC (SEQ ID
NO：20)
TGCTCTCGAGCTTCACGCCGCTGATCTCCTCGCGGTT
optiHAF10- CAGGCGGGCCTCCTCGCTGTACTGGGGGTAGTCGTA
Reverse(101) GGTGCCGTTCTTCACGCTCTCCATGC (SEQ ID
NO：21)
GTGAAGCTCGAGAGCATGGGCACCTACCAGATCCTG
optiHAF11- AGCATCTACAGCACCGTGGCCAGCAGCCTGGCCCTG
Forward(107) GCCATCATGGTGGCCGGCCTGAGCCTGTGGATGTG
(SEQ ID NO：22)
AGTAGACACAAGGGTGTTTTTAACTACAATCTGAACT
optiHAF11- CACAAATTTAGATGCAGATGCGGCACTGCAGGCTGC
Reverse(101) CGTTGCTGCACATCCACAGGCTCAGGCC(SEQ ID
NO：23)
另外，分别根据片段optiHAF2-optiHAF11的2个寡核苷酸片段序列，同时参照片段optiHAF2-optiHAF11的序列，用Oligo 6.0软件(Wojciech&Piotr Rychlik Copyright，Version 6.67)设计用于OE-PCR扩增的上下游引物，片段optiHAF2的两个引物为optiHAF2-Forward-5’和optiHAF2-Reverse-5’；片段optiHAF3的两个引物为optiHAF3-Forward-5’和optiHAF3-Reverse-5’；片段optiHAF4的两个引物为optiHAF4-Forward-5’和optiHAF4-Reverse-5’；片段optiHAF5的两个引物为optiHAF5-Forward-5’和optiHAF5-Reverse-5’；片段optiHAF6的两个引物为optiHAF6-Forward-5’和optiHAF6-Reverse-5’；片段optiHAF7的两个引物为optiHAF7-Forward-5’和optiHAF7-Reverse-5’；片段optiHAF8的两个引物为optiHAF8-Forward-5’和optiHAF8-Reverse-5’；片段optiHAF9的两个引物为optiHAF9-Forward-5’和optiHAF9-Reverse-5’；片段optiHAF10的两个引物为optiHAF10-Forward-5’和optiHAF10-Reverse-5’；片段optiHAF11的两个引物为optiHAF11-Forward-5’和optiHAF11-Reverse-5’。对optiHAF1-Forward(101)片段只设计其上游引物optiHAF1-Forward-5’，所有引物均送由Sigma公司直接合成(见附表3)。
附表3 用于OE-PCR的引物
| 引物名称 | 引物序列 |
| optiHAF1-Forward-5’ | AGCGAAAGCAGGGGTCCAATCTGTCA(SEQ ID NO：24) |
| optiHAF2-Forward-5’ | CCAGATCTGCATCGGCTACCACGCCA(SEQ ID NO：25) |
| optiHAF2-Reverse-5’ | GCCACGCTGCAGTCGCGCAGGATCA(SEQID NO：26) |
| optiHAF3-Forward-5’ | GACTGCAGCGTGGCCGGCTGGCTGCTG(SEQ ID NO：27) |
| optiHAF3-Reverse-5’ | CCAGCTGCTCTTGGGGATGATCTGGATC(SEQ ID NO：28) |
| optiHAF4-Forward-5’ | CAAGAGCAGCTGGAGCAACCACGA(SEQID NO：29) |
| optiHAF4-Reverse-5’ | TGCCCCACAGCACCAGCAGGTCCTC(SEQID NO：30) |
| optiHAF5-Forward-5’ | AGGAGGACCTGCTGGTGCTGTGG(SEQ IDNO：31) |
| optiHAF5-Reverse-5’ | CTCCATGCGGCCGCTCTGCCCGTTC(SEQID NO：32) |
| optiHAF6-Forward-5’ | GAGCGGCCGCATGGAGTTCTTCTGGAC(SEQID NO：33) |
| optiHAF6-Reverse-5’ | GGCATGCTGCTGTTGATGGCGCCCATG(SEQID NO：34) |
| optiHAF7-Forward-5’ | CAGCATGCCCTTCCACAACATCCAC(SEQ |
| ID NO：35) | |
| optiHAF7-Reverse-5’ | GTGGTACCCGTACCAGCCGTCCACCA(SEQ ID NO：36) |
| optiHAF8-Forward-5’ | GGTACGGGTACCACCACAGCAACGA(SEQID NO：37) |
| optiHAF8-Reverse-5’ | GCGGCGGCTCTTCCTCCAGGTTGT(SEQID NO：38) |
| optiHAF9-Forward-5’ | CAACCTGCTCTTCGGAGCGCCGCAT(SEQID NO：39) |
| optiHAF9-Reverse-5’ | GCGCAGCTGCAGGCGCACCTTGTC(SEQID NO：40) |
| optiHAF10-Forward-5’ | GTGCGCCTGCAGCTGCGCGACAAC(SEQID NO：41) |
| optiHAF10-Reverse-5’ | TGCTCTCGAGCTTCACGCCGCTG(SEQ IDNO：42) |
| optiHAF11-Forward-5’ | GTGAAGCTCGAGAGCATGGGCAC(SEQ IDNO：43) |
| optiHAF11-Reverse-5’ | AGTAGACACAAGGGTGTTTTTAACTAC(SEQ ID NO：44) |
实施例3.optiHA基因11个片段optiHAF1-optiHAF11的OE-PCR扩增、克隆及测序鉴定
11个片段optiHAF1-optiHAF11分别用PlatihumPfx Taq DNA聚合酶扩增。首先在PCR管中依次加入10X Pfx Taq聚合酶缓冲液10ul、2.5mmoldNTPs 4ul、Pfx Taq DNA聚合酶1ul(1-2U)以及对应的正反向寡核苷酸片段(对于optiHAF1只加入正向寡核苷酸片段)各10ul，灭菌去离子水补足100ul后，95℃5min，48℃～55℃退火1min，72℃5min，5个循环后再加入20pmol对应于各个寡核苷酸片段的上、下游引物(附表3)各1ul在PTC-100基因扩增仪(M.J.Research.inc)上扩增(对于optiHAF1只加入上游引物)。OE-PCR的反应条件是48℃1min、72℃1.5min，30个循环，最后72℃ 5min延伸，取5ul PCR产物在1％琼脂糖凝胶上电泳检查扩增结果，片段optiHAF1-optiHAF11的大小依次分别为：88bp、162bp、204bp、162bp、164bp、201bp、201bp、155bp、157bp、168bp和182bp(见图2)。
将11个片段的PCR产物按照胶回收(小量)试剂盒说明书操作步骤回收，之后分别克隆于pBluscript载体的SmaI位点，转化受体菌E.coliJM109后分别随机选取4个克隆，按照质粒(小量)抽提试剂盒说明书提取质粒，溶于50μL的灭菌去离子水中，用CEQTM DTCS-QUICK Star Kit进行质粒DNA模板的测序前处理，具体步骤如下：分别将1μL的11个片段的克隆质粒和6μL灭菌去离子水混合，96℃加热1-3分钟，自然凉到室温后加入T3引物(T3：ATTAACCCTCACTAAAGGGAA，SEQ ID NO：45)1μl(5pmol/μl)和DTCS Quick Start Master Mix 2μl，混匀后进行测序PCR反应，PCR反应程序为：96℃20sec、50℃20sec、60℃4min，35个循环，最后保持在4℃。PCR反应完后加入2.5ul的反应终止液(3M PH5.2的NaOAC 2ul，100mM PH8.0的EDTA 2ul，20mg/ml的Glycogen 1ul，即用即配)终止反应，然后加入30ul 95％冰乙醇沉淀DNA，混匀，4℃、12000rpm立即离心20min。小心倒掉上清后，用100～200ul 70％冰乙醇两次洗脱残留的盐，4℃、12000rpm离心5min。真空抽干或室温晾干DNA后，用30ul甲酰胺充分溶解DNA沉淀，小心加入CEQ样品板孔内，最后加一滴石蜡油到样品板孔内。随即采用双脱氧链终止法在BechmanCEQ8000测序仪(Bechman，America)上对克隆重组质粒进行测序，之后利用DNASTAR软件(DNASTAR.Inc.)将得到的序列进行分析，结果所挑取的每个片段的4个克隆中，分别有1-2个为阳性克隆，阳性克隆依次命名为pBluscript-optiHAF1至pBluscript-optiHAF11。
实施例4.基因optiHA的连接构建策略
将实施例3中测序验证好的11个片段的阳性质粒pBluscript-optiHAF1至pBluscript-optiHAF11根据实施例1中所选定的单酶切位点按以下方法和顺序，使用常规技术依次酶切连接：(1)将pBluscript-optiHAF1用XbaI酶切后用klenow酶补平、BglII酶切后回收大小为2700bp的片段，将pBluscript-optiHAF2用BglII和EcoRV双酶切后回收大小为160bp的片段，然后将所回收的两个片段相连接成含有optiHAF1与optiHAF2的质粒pBluscript-optiHAF1-2，将pBluscript-optiHAF1-2与pBluscript-optiHAF3分别用PstI和HindIII酶切后连接成pBluscript-optiHAF1-2-3，再将用SacI酶切、T4 DNA聚合酶补平和EcoRI处理后的pBluscript-optiHAF1-2-3与用KpnI酶切、klenow酶补平和EcoRI处理后的载体pCI相连接而得到pCI-optiHAF1-2-3；(2)将pBluscript-optiHAF5与pBluscript-optiHAF6分别用NotI和HindIII酶切后连接成pBluscript-optiHAF5-6，将pBluscript-optiHAF5-6与pBluscript-optiHAF7分别用SphI和BamHI酶切后连接成pBluscript-optiHAF5-6-7，再将pBluscript-optiHAF5-6-7与pBluscript-optiHAF4分别用EcoO109I和BamHI酶切后连接成pBluscript-optiHAF4-5-6-7；(3)先将pBluscript-optiHAF9和载体pCI分别用XbaI和EcoRI处理后连接成pCIoptiHAF9，将pCI-optiHAF9与pBluscript-optiHAF8分别用SapI和EcoRI酶切后连接成pCIoptiHAF8-9，将pBluscript-optiHAF10与pBluscript-optiHAF11分别用BamHI和EcoRI酶切后连接成pBluscript-optiHAF10-11，再将pCI-optiHAF8-9与pBluscript-optiHAF10-11分别用PstI和XbaI酶切后连接成pCI-optiHAF8-9-10-11；(4)将pCI-optiHAF1-2-3与pBluscript-optiHAF4-5-6-7分别用PvuII和XbaI酶切后连接成pCI-optiHAF1-2-3-4-5-6-7，再将pCI-optiHAF8-9-10-11与pCI-optiHAF1-2-3-4-5-6-7分别用KpnI和XbaI酶切后连接成含有全长的optiHA基因的质粒pCI-optiHAFull(连接策略见图3)。
实施例5.基因optiHA的PCR鉴定及酶切分析
含有optiHA基因的质粒pCIoptiHAFull用pCI上的上游引物：pCI-up：5’-ACTTAATACGACTCACTATA-3’(SEQ ID NO：46)；基因的下游引物：5‘-CCACTCTAGAGTGTTTTTAACTACAA-3’(SEQ ID NO：47)进行PCR鉴定，PCR条件为：94℃ 5min、94℃ 1min、50℃ 1min、72℃ 1.5min，35个循环，最后72℃ 10min延伸，并将PCR产物进行琼脂糖凝胶(1％)电泳，结果得到了长约1.7kb的条带，与A/Goose/Guangdong/1/96(H5N1)[GD/1/96(H5N1)]HA基因的片段大小一致(见图4)。按照限制性内切酶的说明书，将质粒pCIoptiHAFull进行酶切处理，并将酶切产物进行琼脂糖凝胶(1％)电泳，结果，用PvuII酶切时得到了1.0Kb和4.8kb的两条带、用XhoI酶切得到了1.6kb和4.2kb的条带、用NotI酶切得到了1.2kb和4.6kb的条带、用EcoRI酶切时得到了线性化的5.8kb的片段、用BamHI酶切时得到了300bp和5.5kb的条带，所得结果均与对所书写的optiHA基因序列(SEQ ID NO：1)的分析结果一致(见图5)。
实施例6.基因optiHA的测序验证
将含有optiHA基因的质粒pCIoptiHAFull按照质粒(小量)抽提试剂盒说明书提取，用CEQTM DTCS-QUICK Star Kit进行质粒DNA模板的测序前处理，具体步骤如下：在5个PCR管中分别将1ul的质粒pCIoptiHAFullDNA模板和6ul灭菌去离子水混合，96℃加热1-3分钟，自然凉到室温后依次分别加入pCI-up：5’-ACTTAATACGACTCACTATA-3’(SEQ IDNO：48)；opt-w1p：5’-GCACCAACCACTTCGAGA-3’(SEQ ID NO：49)；opt-w1f：5‘-GCTACCACGCCAACAACAGC-3’(SEQ ID NO：50)；opt-w2p：5‘-GAAGAGCGAGCTGGAGTAC-3’(SEQ ID NO：51)；opt-w3p：5‘-GCAGATGGAGGACGGCTTCCT-3’(SEQ ID NO：52)；pCI-lower：5‘-TGCAGCTTATAATGGTTACA-3’(SEQ ID NO：53)1μl(5pmol/μl)和DTCS Quick Start Master Mix 2μl，混匀后进行测序PCR反应，PCR反应程序为：96℃20sec、50℃20sec、60℃4min，35个循环，最后保持在4℃。PCR反应完后加入2.5ul的反应终止液(3M PH5.2的NaOAC 2ul，100mM PH8.0的EDTA 2ul，20mg/ml的Glycogen 1ul，即用即配)终止反应，然后加入30ul 95％冰乙醇沉淀DNA，混匀，4℃、12000rpm立即离心20min。小心倒掉上清后，用100～200ul 70％冰乙醇两次洗脱残留的盐，4℃、12000rpm离心5min。真空抽干或室温晾干DNA后，用30ul甲酰胺充分溶解DNA沉淀，每个PCR管中的样品分别小心加入到CEQ样品板孔内，最后加一滴石蜡油到样品板孔内。双脱氧链终止法在BechmanCEQ8000测序仪上对优化的基因optiHA进行测序验证，经过测序，所构建的optiHA基因与所书写的optiHA基因序列(SEQ ID NO：1)完全一致，基因总长为1779bp，阅读框架含核苷酸1707bp，共编码568个氨基酸(SEQ ID NO：2)。
实施例7.基因optiHA表达质粒pCIoptiHA的构建
用特异性引物(上游：5‘-TCAAAATGGAGAAGATCGTGCT-3’(SEQID NO：54)，下游：5‘-CCACTCTAGAGTGTTTTTAACTACAA-3’(SEQ IDNO：55))从质粒pCIoptiHAFull中用Pfx Taq DNA聚合酶扩增仅含开放阅读框架(ORF)的optiHA基因。在PCR管中依次加入10X Pfx Taq聚合酶缓冲液10ul、2.5mmol dNTP s4ul、Pfx Taq DNA聚合酶1ul(1-2U)、质粒pCIoptiHAF1ul以及对应的上下游引物1ul，灭菌去离子水补足100ul，扩增条件为：94℃5min、94℃ 1min、58℃1min、72℃ 1.5min，35个循环，最后72℃10min延伸，将PCR产物用XbaI彻底酶切后定向克隆到用KpnI酶切后用klenow酶补平、再用XbaI酶切处理的质粒载体pCI中，得到质粒pCIoptiHA，将构建好的质粒pCIoptiHA进行酶切鉴定，并将酶切产物进行琼脂糖凝胶(1％)电泳，结果，质粒pCIoptiHA用EcoRI酶切时得到了线性化的5.8kb的片段，用PstI酶切时得到了1.2Kb、530bp、300bp和3.87kb的条带、用BglII酶切得到了1.1kb和4.7kb的条带，所得结果均与对所构建的optiHA基因序列(SEQ ID NO：1)的分析结果一致(见图6)。
实施例8.基因optiHA在293T细胞中的表达
转染用质粒pCIoptiHA和pCIHA(作为对照，将含有禽流感病毒A/Goose/GuangDong/1/96(H5N1)[GD/1/96(H5N1)]HA基因的克隆质粒pBH5(陈化兰，于康震，步志高。一株鹅源高致病力禽流感病毒分离株血凝素基因的分析。中国农业科学，1999，32(2)：87-92.))经EcoRI酶切、klenow酶补平及SalI酶切后切下HA基因片段，插入SalI和SmaI酶切的CMV启动子表达载体pCI而构建出表达质粒pCIHA)按照PerfectprepPlasmid Mini操作步骤进行提取。293T细胞(人胚胎肾细胞)先以含10％胎牛血清的DMEM培养液培养于细胞培养瓶中，连续传代5代以上后传于六孔板中，置于37℃5％CO2培养箱中培养。待293T细胞长至60％～80％铺满时，按LipofectamineTM2000试剂盒使用说明书进行转染，转染后通过间接免疫荧光和Western blot检测其表达的HA蛋白。
8.1 间接免疫荧光检测
质粒pCIoptiHA(pCIHA作为对照)转染293T细胞后24h、48h时，进行间接免疫荧光检测，同时设未转染质粒的293T细胞阴性对照。用70％乙醇固定20min-30min后，自然干燥。用PBS(0.01mol/L，pH7.4)洗涤后，取H5亚型禽流感多克隆阳性血清用PBS(0.01mol/L，pH7.4)20倍稀释，滴加在6孔板中央选定的区域内，37℃作用45min。用PBS洗涤三次，每次5min，洗后用去离子水再冲一次，自然干燥。取FIFC标记的兔抗鸡IgG(FITC-IgG)用含伊文斯蓝(1/104)的PBS(0.01mol/L，pH7.4)100倍稀释，滴加在6孔板中央选定的区域内，37℃作用45min。之后用PBS(0.01mol/L，pH7.4)洗涤三次，用去离子水脱盐后，自然干燥。滴加碱性甘油(pH9.8)，于荧光显微镜(Leica DMIRE2)下观察。结果，质粒pCIoptiHA和pCIHA在转染293T细胞后，经过间接免疫荧光染色，表达的HA蛋白产物主要分布于细胞膜上，并在荧光显微镜下呈现出亮绿色的特殊荧光(如图7、8、9和10中箭头所示)。2种质粒转染293T细胞后24h均即可表达H5亚型HA蛋白(见图7和图8)，在转染后48h，能表达HA蛋白的细胞数明显比24h的多(见图9，图10)。在转染后24h和48h，优化的基因optiHA的表达水平均明显高于野生型HA基因的表达水平(见图8，图10)。未转染质粒的293T细胞对照(见图11)未见有特异的荧光表达产物。
8.2 Western blot检测
用质粒转染后48h的293T细胞及H5亚型禽流感病毒GD/1/96(H5N1)裂解后制备的样品进行SDS-PAGE电泳，之后转移到硝酸纤维素(NC)膜上用H5亚型禽流感多克隆阳性血清进行Western blot检测2种质粒所表达的AIV HA蛋白。
8.2.1 SDS-PAGE电泳
样品制备
在质粒pCIoptiHA(pCIHA作为对照)转染48h后收获293T细胞，用PBS(pH7.4)洗三遍，加入与细胞收集液等倍体积的预热的2×SDS电泳缓冲液裂解细胞，收集到1.5ml微量离心管中，置于沸水中煮10min使蛋白样品充分变性即可。H5亚型禽流感病毒GD/1/96(H5N1)作为阳性对照，在SPF鸡胚中繁殖后收集尿囊液，其SDS-PAGE电泳样品的制备过程同上。
凝胶灌制及电泳
分别配制10％分离胶，5％的浓缩胶，按《分子克隆实验指南》(第二版)(美J.萨姆布鲁克等编著，金冬雁等译，2002，科学出版社)所述进行灌制，同时灌制两块凝胶。待其聚合后，用微量移液器分别加入10ul质粒pCIoptiHA转染后48h的293T细胞、质粒pCIHA转染后48h的293T细胞、H5亚型禽流感病毒GD/1/96(H5N1)裂解后制备的样品及293T细胞，同时加蛋白电泳分子量标准(SM0431，晶美生物工程有限公司)作对照，在BIO-RAD垂直电泳槽上先80V电泳，待样品进入分离胶后调整电压为120V，待染料移至凝胶底部时停止电泳。将其中一凝胶置于0.25％考马斯亮蓝染色液中(0.25％R250，40％甲醇，10％乙醇)2小时，用脱色液(40％甲醇，10％乙酸)脱色，结果转染质粒的293T细胞的条带与293T细胞对照的条带一致，而流感病毒对照的不同的蛋白条带清晰可见(见图12)。另一凝胶置于转印装置，用于Western-blot检测。
8.2.2 Western blot检测
电转移
将6张Whatman滤纸和NC膜剪成与凝胶同样大小，在转移缓冲液(48mmol/L Tris，39mmol/L甘氨酸，0.037％SDS)中浸泡5min，按3张滤纸、NC膜、凝胶、3张滤纸的顺序依次将起叠放在一起，放置每一层均要除去它们之间的气泡，按NC膜侧靠阳极、凝胶侧靠阴极将其放入转印槽中，在TRANS-BLOT SD(Semi-dry Transfer Cell BIO-RAD)转印装置22V电转移45min，取出NC膜，作好标记，夹于滤纸之间室温干燥30min。然后22V电转45min(纤维素膜侧靠正极，胶侧靠负极)。转印完毕，取出NC膜用铅笔在膜上作好标记。夹于两层滤纸之间，室温干燥30min。
免疫检测
将NC膜放在一平皿中，加封闭液(0.02％叠氮钠，1％(w/v)BSA溶于PBS)，浸过膜即可，室温下轻振2-3h。然后用PBS洗3次，加H5亚型禽流感多克隆阳性血清作为一抗(用PBS 1∶100稀释)4℃作用4-5h或过夜，再用PBS洗3次，Buffer I(150mmol/L NaCl，50mmol/L Tris.ClpH7.5)洗10min，加入辣根过氧化物酶标记的兔抗鸡IgG(HRPO-IgG)(用含1％BSA的Buffer I作1∶1000稀释)37℃作用2h。用Buffer I洗3次，Buffer II(10mmol/L Tris.Cl pH7.5，150mmol/L NaCl)洗一次，吸净余液，用DAB显色试剂盒(包括DAB×20倍浓缩液、H2O2×20倍浓缩液、TBS×20倍浓缩缓冲液)显色3分钟。显色完毕，加0.5mM EDTA(pH8.0)终止显色。
结果2种质粒均可检测出分子量约为74Kda的蛋白，优化的基因optiHA比野生型HA基因的表达水平更高(见图13)。
实施例9.表达optiHA的质粒pCIoptiHA对SPF鸡的免疫保护试验
9.1 免疫保护试验
将3周龄SPF鸡随机分为5组，4或5只/组。第1组用pCIHA 100μg免疫，经腿部分两点肌肉注射200μl 0.5μg/μl的pCIHA。第2组为pCIHA10μg免疫组，经腿部分两点肌肉注射200μl 0.05μg/μl的pCIHA；第3组为pCIoptiHA 100μg免疫，免疫方法同第1组，第4组为pCIoptiHA 10μg免疫，免疫方法同第2组。第5组为对照组，注射200μl PBS溶液。两种DNA疫苗质粒在单次免疫后4周分别用100LD50的HPAIVA/Goose/Guangdong/1/96(H5N1)[GD/1/96(H5N1)]经鼻腔途径感染，0.1ml/只，攻毒后2周内每天观察记录鸡只的发病和死亡情况，并根据死亡数统计死亡率。对每组鸡于免疫后和攻毒后每周采集其翅静脉血，分离血清，分别采用血凝抑制试验(HI)(高致病性禽流感诊断技术(GB/T/18936-2003))、琼脂扩散试验(AGP)(高致病性禽流感诊断技术(GB/T/18936-2003))测定其相应HI抗体和AGP抗体滴度。在攻毒后第三天、第五天和第七天采集每组鸡的泄殖腔拭子和喉头拭子，将每个拭子用PBS溶液从10-1至10-6稀释度作10倍连续稀释，每个稀释度经尿囊腔接种3枚9～11日龄鸡胚，0.1ml/枚，48h收集尿囊液，微量血凝法(高致病性禽流感诊断技术(GB/T/18936-2003))检测病毒，并根据其不同稀释度的血凝结果用Reed & Muench法(殷震等著，动物病毒学，第二版，1997，科学出版社，329.)计算其病毒分离的滴度。
9.2 试验结果
9.2.1 免疫及强毒攻击后的HI抗体反应
pCIHA和pCIoptiHA分别以100μg和10μg剂量免疫一次后，免疫鸡的抗体转阳情况以及抗体水平明显不同。100μg pCIHA在免疫后3周只有1只鸡可检测到抗体，100μg pCIoptiHA免疫后2周有2只鸡检测到抗体，免疫后4周仅有1只鸡没有抗体，10μg pCIHA免疫后没有检测到抗体，而10μg pCIoptiHA免疫后2周有2只鸡检测到抗体；pCIoptiHA以100μg和10μg单次免疫后诱导的HI抗体水平远远高于相同剂量pCIHA免疫后的HI抗体水平(见表4)。
表4.pCIHA和pCIoptiHA单次免疫后HI抗体转阳情况及免疫和攻毒后
HI抗体(log2)的动态变化
| 组别 | 免疫原 | 剂量 | 免疫后(周) | 攻毒后(周) | ||||||||
| 1 | 2 | 3 | 4* | 1 | 2 | |||||||
| 1 | pCIHA | 100μg | 0/4 | 0 | 0/4 | 0 | 1/4 | 0.6±1.3 | 1/4 | 1.0±2.0 | 2.3±1.5 | 5.8±1.1 |
| 2 | pCIHA | 10μg | 0/5 | 0 | 0/5 | 0 | 0/5 | 0 | 0/5 | 0 | - | - |
| 3 | pCIoptiHA | 100μg | 0/4 | 0 | 2/4 | 2.0±2.8 | 3/4 | 3.1±2.9 | 3/4 | 3.5±3.5 | 4.0±2.0 | 7.0±1.7 |
| 4 | pCIoptiHA | 10μg | 0/4 | 0 | 2/4 | 1.8±2.4 | 2/4 | 2.4±2.8 | 2/4 | 1.9±2.6 | 3.0±3.0 | 5.0±2.6 |
| 5 | 对照 | - | 0/5 | 0 | 0/5 | 0 | 0/5 | 0 | 0/5 | 0 | - | - |
注：抗体转阳表示方法为HI抗体转阳鸡数/免疫鸡总数，-表示在攻毒后鸡群已全部死亡。*免疫后第四周攻毒。
9.2.2 发病和死亡情况统计及AGP抗体检测
GD/1/96(H5N1)HPAIV攻击后，所有对照鸡在攻毒后2至7天内全部死亡。pCIoptiHA 100μg一次免疫后有1只鸡发病并死亡，其死亡保护率为75％，pCIHA 100μg单免组在病毒攻击后11天内死亡2只鸡；pCIoptiHA10μg的保护率为75％，pCIHA 10μg单免组在病毒攻击后的7天之内即死亡4只鸡，另外1只鸡在攻毒后第11天死亡。免疫后血清AGP抗体检测均为阴性；在攻毒后2周时，pCIHA 100μg单免组中有1只鸡AGP抗体为阳性，其它所有存活鸡的AGP抗体检测均为阴性(见表5)。
表5.pCIHA和pCIoptiHA免疫鸡用HPAIV H5N1攻击后发病与死亡情况
及AGP抗体检测结果
| 组别 | 免疫原 | 剂量 | 发病数/鸡总数 | 死亡数/鸡总数 | 保护率(存活/总数) | 免疫后AGP抗体(周) | HPAIV攻击后AGP抗体(周) | ||||
| 1 | 2 | 3 | 4* | 1 | 2 | ||||||
| 1 | pCIHA | 100μg | 2/4 | 2/4 | 2/4 | - | - | - | - | 0/2 | 1/2 |
| 2 | pCIHA | 10μg | 5/5 | 5/5 | 0/5 | - | - | - | - | × | × |
| 3 | pCIoptiHA | 100μg | 1/4 | 1/4 | 3/4 | - | - | - | - | 0/3 | 0/3 |
| 4 | pCIoptiHA | 10μg | 1/4 | 1/4 | 3/4 | - | - | - | - | 0/3 | 0/3 |
| 5 | 对照 | - | 5/5 | 5/5 | 0/5 | - | - | - | - | × | × |
注：*免疫后第四周攻毒；-代表所有实验鸡AGP抗体检测为阴性；GD/1/96(H5N1)HPAIV攻击后AGP抗体表示方法为：斜线‘/’的上面和下面分别为AGP抗体阳性鸡数和当时存活鸡数；×表示鸡在攻毒一周后已全部死亡。
9.2.3 病毒分离滴定结果
将采集的泻殖腔和喉头拭子接种鸡胚进行病毒分离和滴定，结果，100μg pCIoptiHA和pCIHA单次免疫形成不完全免疫保护；pCIoptiHA以10μg单次免疫也形成不完全免疫保护；pCIHA以10μg单次免疫几乎不能形成免疫保护，GD/1/96(H5N1)HPAIV攻击后的3天、5天和7天均可分离到不同滴度的病毒(见表6)。
表6.pCIHA和pCIoptiHA单次免疫鸡用H5N1攻击后病毒分离及滴定结
果
| 组别 | 免疫原 | 剂量 | 拭子 | 攻毒后(天) | |||||
| 3 | 5 | 7 | |||||||
| 排毒 | 滴度 | 排毒 | 滴度 | 排毒 | 滴度 | ||||
| 1 | pCIHA | 100μg | 喉头 | 1/3 | 0.97 | 1/3 | 0.9 | 0/3 | ＜0.9 |
| 泄殖腔 | 2/3 | 1.58 | 0/3 | ＜0.9 | 0/3 | ＜0.9 | |||
| 2 | pCIHA | 10μg | 喉头 | 2/5 | 1.7 | 0/3 | ＜0.9 | 1/1 | 0.9 |
| 泄殖腔 | 3/5 | 1.1 | 1/3 | 0.9 | 0/1 | ＜0.9 | |||
| 3 | pCIoptiHA | 100μg | 喉头 | 1/4 | 2.5 | 0/3 | ＜0.9 | 0/3 | ＜0.9 |
| 泄殖腔 | 1/4 | 0.98 | 0/3 | ＜0.9 | 0/3 | ＜0.9 | |||
| 4 | pCIoptiHA | 10μg | 喉头 | 0/3 | ＜0.9 | 0/3 | ＜0.9 | 0/3 | ＜0.9 |
| 泄殖腔 | 0/3 | ＜0.9 | 0/3 | ＜0.9 | 0/3 | ＜0.9 | |||
| 5 | Control | --- | 喉头 | 2/3 | 1.2 | 0/1 | ＜0.9 | - | - |
| 泄殖腔 | 3/3 | 1.47 | 0/1 | ＜0.9 | - | - | |||
注：斜线‘/’的上面和下面分别为病毒分离阳性鸡数和当时存活鸡数，拭子原液为阳性时，病毒滴定结果定义为0.9。
9.3 结论
试验结果表明，HA基因密码子优化而得到的optiHA基因可显著提高H5亚型禽流感DNA疫苗诱导的保护性抗体免疫反应水平，提高免疫原性，增强免疫保护效果。
I040426-序列表
SEQUENCE LISTING
<110>中国农业科学院哈尔滨兽医研究所
<120>编码H5亚型禽流感病毒血凝素蛋白的基因及其应用
<130>I040426
<160>55
<170>PatentIn version 3.1
<210>1
<211>1779
<212>DNA
<213>人工合成
<220>
<221>CDS
<222>(29)..(1732)
<223>
<400>1
agcgaaagca ggggtccaat ctgtcaaa atg gag aag atc gtg ctg ctg ctg 52
Met Glu Lys Ile Val Leu Leu Leu
1 5
gcc atc gtg agc ctg gtg aag agc gac cag atc tgc atc ggc tac cac 100
Ala Ile Val Ser Leu Val Lys Ser Asp Gln Ile Cys Ile Gly Tyr His
10 15 20
gcc aac aac agc acc gag cag gtg gac acc atc atg gag aag aac gtg 148
Ala Asn Asn Ser Thr Glu Gln Val Asp Thr Ile Met Glu Lys Asn Val
25 30 35 40
acc gtg acc cac gcc cag gac atc ctg gag aag acc cac aac ggc aag 196
Thr Val Thr His Ala Gln Asp Ile Leu Glu Lys Thr His Asn Gly Lys
45 50 55
ctg tgc gac ctg aac ggc gtg aag ccc ctg atc ctg cgc gac tgc agc 244
Leu Cys Asp Leu Asn Gly Val Lys Pro Leu Ile Leu Arg Asp Cys Ser
60 65 70
gtg gcc ggc tgg ctg ctg ggc aac ccc atg tgc gac gag ttc atc aac 292
Val Ala Gly Trp Leu Leu Gly Asn Pro Met Cys Asp Glu Phe Ile Asn
75 80 85
gtg ccc gag tgg agc tac atc gtg gag aag gcc agc ccc gcc aac gac 340
Val Pro Glu Trp Ser Tyr Ile Val Glu Lys Ala Ser Pro Ala Asn Asp
90 95 100
ctg tgc tac ccc ggc gac ttc aac gac tac gag gag ctg aag cac ctg 388
Leu Cys Tyr Pro Gly Asp Phe Asn Asp Tyr Glu Glu Leu Lys His Leu
I040426-序列表
105 110 115 120
ctg agc cgc acc aac cac ttc gag aag atc cag atc atc ccc aag agc 436
Leu Ser Arg Thr Asn His Phe Glu Lys Ile Gln Ile Ile Pro Lys Ser
125 130 135
agc tgg agc aac cac gac gcc agc agc ggc gtg agc agc gcc tgc ccc 484
Ser Trp Ser Asn His Asp Ala Ser Ser Gly Val Ser Ser Ala Cys Pro
140 145 150
tac cac ggc cgc agc agc ttc ttc cgc aac gtg gtg tgg ctg atc aag 532
Tyr His Gly Arg Ser Ser Phe Phe Arg Asn Val Val Trp Leu Ile Lys
155 160 165
aag aac agc gcc tac ccc acc atc aag cgc agc tac aac aac acc aac 580
Lys Asn Ser Ala Tyr Pro Thr Ile Lys Arg Ser Tyr Asn Asn Thr Asn
170 175 180
cag gag gac ctg ctg gtg ctg tgg ggc atc cac cac ccc aac gac gcc 628
Gln Glu Asp Leu Leu Val Leu Trp Gly Ile His His Pro Asn Asp Ala
185 190 195 200
gcc gag cag acc aag ctg tac cag aac ccc acc acc tac atc agc gtg 676
Ala Glu Gln Thr Lys Leu Tyr Gln Asn Pro Thr Thr Tyr Ile Ser Val
205 210 215
ggc acc agc acc ctg aac cag cgc ctg gtg ccc gag atc gcc acc cgc 724
Gly Thr Ser Thr Leu Asn Gln Arg Leu Val Pro Glu Ile Ala Thr Arg
220 225 230
ccc aag gtg aac ggg cag agc ggc cgc atg gag ttc ttc tgg acc atc 772
Pro Lys Val Asn Gly Gln Ser Gly Arg Met Glu Phe Phe Trp Thr Ile
235 240 245
ctg aag ccc aac gac gcc atc aac ttc gag agc aac ggc aac ttc atc 820
Leu Lys Pro Asn Asp Ala Ile Asn Phe Glu Ser Asn Gly Asn Phe Ile
250 255 260
gcc ccc gag tac gcc tac aag atc gtg aag aag ggc gac agc gcc atc 868
Ala Pro Glu Tyr Ala Tyr Lys Ile Val Lys Lys Gly Asp Ser Ala Ile
265 270 275 280
atg aag agc gag ctg gag tac ggc aac tgc aac acc aag tgc cag acc 916
Met Lys Ser Glu Leu Glu Tyr Gly Asn Cys Asn Thr Lys Cys Gln Thr
285 290 295
ccc atg ggc gcc atc aac agc agc atg ccc ttc cac aac atc cac ccc 964
Pro Met Gly Ala Ile Asn Ser Ser Met Pro Phe His Asn Ile His Pro
300 305 310
ctg acc atc ggc gag tgc ccc aag tac gtg aag agc aac cgc ctg gtg 1012
Leu Thr Ile Gly Glu Cys Pro Lys Tyr Val Lys Ser Asn Arg Leu Val
315 320 325
ctg gcc acc ggc ctg cgc aac acc ccc cag cgc gag cgc cgc cgc aag 1060
Leu Ala Thr Gly Leu Arg Asn Thr Pro Gln Arg Glu Arg Arg Arg Lys
330 335 340
aag cgc ggc ctg ttc ggc gcc atc gcc ggc ttc atc gag ggc ggc tgg 1108
Lys Arg Gly Leu Phe Gly Ala Ile Ala Gly Phe Ile Glu Gly Gly Trp
345 350 355 360
cag ggc atg gtg gac ggc tgg tac ggg tac cac cac agc aac gag cag 1156
Gln Gly Met Val Asp Gly Trp Tyr Gly Tyr His His Ser Asn Glu Gln
365 370 375
ggc agc ggc tac gcc gcc gac aag gag agc acc cag aag gcc atc gac 1204
Gly Ser Gly Tyr Ala Ala Asp Lys Glu Ser Thr Gln Lys Ala Ile Asp
380 385 390
ggc gtg acc aac aag gtg aac agc atc atc gac aag atg aac acc cag 1252
Gly Val Thr Asn Lys Val Asn Ser Ile Ile Asp Lys Met Asn Thr Gln
395 400 405
ttc gag gcc gtg ggc cgc gag ttc aac aac ctg gag cgc cgc atc gag 1300
I040426-序列表
Phe Glu Ala Val Gly Arg Glu Phe Asn Asn Leu Glu Arg Arg Ile Glu
410 415 420
aac ctg aac aag cag atg gag gac ggc ttc ctg gac gtg tgg acc tac 1348
Asn Leu Asn Lys Gln Met Glu Asp Gly Phe Leu Asp Val Trp Thr Tyr
425 430 435 440
aac gcc gag ctg ctg gtg ctg atg gag aac gag cgc acc ctg gac ttc 1396
Asn Ala Glu Leu Leu Val Leu Met Glu Asn Glu Arg Thr Leu Asp Phe
445 450 455
cac gac agc aac gtg aag aac ctg tac gac aag gtg cgc ctg cag ctg 1444
His Asp Ser Asn Val Lys Asn Leu Tyr Asp Lys Val Arg Leu Gln Leu
460 465 470
cgc gac aac gcc aag gag ctg ggc aac ggc tgc ttc gag ttc tac cac 1492
Arg Asp Asn Ala Lys Glu Leu Gly Asn Gly Cys Phe Glu Phe Tyr His
475 480 485
aag tgc gac aac gag tgc atg gag agc gtg aag aac ggc acc tac gac 1540
Lys Cys Asp Asn Glu Cys Met Glu Ser Val Lys Asn Gly Thr Tyr Asp
490 495 500
tac ccc cag tac agc gag gag gcc cgc ctg aac cgc gag gag atc agc 1588
Tyr Pro Gln Tyr Ser Glu Glu Ala Arg Leu Asn Arg Glu Glu Ile Ser
505 510 515 520
ggc gtg aag ctc gag agc atg ggc acc tac cag atc ctg agc atc tac 1636
Gly Val Lys Leu Glu Ser Met Gly Thr Tyr Gln Ile Leu Ser Ile Tyr
525 530 535
agc acc gtg gcc agc agc ctg gcc ctg gcc atc atg gtg gcc ggc ctg 1684
Ser Thr Val Ala Ser Ser Leu Ala Leu Ala Ile Met Val Ala Gly Leu
540 545 550
agc ctg tgg atg tgc agc aac ggc agc ctg cag tgc cgc atc tgc atc 1732
Ser Leu Trp Met Cys Ser Asn Gly Ser Leu Gln Cys Arg Ile Cys Ile
555 560 565
taaatttgtg agttcagatt gtagttaaaa acacccttgt gtctact 1779
<210>2
<211>568
<212>PRT
<213>人工合成
<400>2
Met Glu Lys Ile Val Leu Leu Leu Ala Ile Val Ser Leu Val Lys Ser
1 5 10 15
Asp Gln Ile Cys Ile Gly Tyr His Ala Asn Asn Ser Thr Glu Gln Val
20 25 30
Asp Thr Ile Met Glu Lys Asn Val Thr Val Thr His Ala Gln Asp Ile
35 40 45
Leu Glu Lys Thr His Asn Gly Lys Leu Cys Asp Leu Asn Gly Val Lys
50 55 60
Pro Leu Ile Leu Arg Asp Cys Ser Val Ala Gly Trp Leu Leu Gly Asn
65 70 75 80
Pro Met Cys Asp Glu Phe Ile Asn Val Pro Glu Trp Ser Tyr Ile Val
I040426-序列表
85 90 95
Glu Lys Ala Ser Pro Ala Asn Asp Leu Cys Tyr Pro Gly Asp Phe Asn
100 105 110
Asp Tyr Glu Glu Leu Lys His Leu Leu Ser Arg Thr Asn His Phe Glu
115 120 125
Lys Ile Gln Ile Ile Pro Lys Ser Ser Trp Ser Asn His Asp Ala Ser
130 135 140
Ser Gly Val Ser Ser Ala Cys Pro Tyr His Gly Arg Ser Ser Phe Phe
145 150 155 160
Arg Asn Val Val Trp Leu Ile Lys Lys Asn Ser Ala Tyr Pro Thr Ile
165 170 175
Lys Arg Ser Tyr Asn Asn Thr Asn Gln Glu Asp Leu Leu Val Leu Trp
180 185 190
Gly Ile His His Pro Asn Asp Ala Ala Glu Gln Thr Lys Leu Tyr Gln
195 200 205
Asn Pro Thr Thr Tyr Ile Ser Val Gly Thr Ser Thr Leu Asn Gln Arg
210 215 220
Leu Val Pro Glu Ile Ala Thr Arg Pro Lys Val Asn Gly Gln Ser Gly
225 230 235 240
Arg Met Glu Phe Phe Trp Thr Ile Leu Lys Pro Asn Asp Ala Ile Asn
245 250 255
Phe Glu Ser Asn Gly Asn Phe Ile Ala Pro Glu Tyr Ala Tyr Lys Ile
260 265 270
Val Lys Lys Gly Asp Ser Ala Ile Met Lys Ser Glu Leu Glu Tyr Gly
275 280 285
Asn Cys Asn Thr Lys Cys Gln Thr Pro Met Gly Ala Ile Asn Ser Ser
290 295 300
Met Pro Phe His Asn Ile His Pro Leu Thr Ile Gly Glu Cys Pro Lys
305 310 315 320
Tyr Val Lys Ser Asn Arg Leu Val Leu Ala Thr Gly Leu Arg Asn Thr
325 330 335
Pro Gln Arg Glu Arg Arg Arg Lys Lys Arg Gly Leu Phe Gly Ala Ile
340 345 350
Ala Gly Phe Ile Glu Gly Gly Trp Gln Gly Met Val Asp Gly Trp Tyr
355 360 365
Gly Tyr His His Ser Asn Glu Gln Gly Ser Gly Tyr Ala Ala Asp Lys
370 375 380
I040426-序列表
Glu Ser Thr Gln Lys Ala Ile Asp Gly Val Thr Asn Lys Val Asn Ser
385 390 395 400
Ile Ile Asp Lys Met Asn Thr Gln Phe Glu Ala Val Gly Arg Glu Phe
405 410 415
Asn Asn Leu Glu Arg Arg Ile Glu Asn Leu Asn Lys Gln Met Glu Asp
420 425 430
Gly Phe Leu Asp Val Trp Thr Tyr Asn Ala Glu Leu Leu Val Leu Met
435 440 445
Glu Asn Glu Arg Thr Leu Asp Phe His Asp Ser Asn Val Lys Asn Leu
450 455 460
Tyr Asp Lys Val Arg Leu Gln Leu Arg Asp Asn Ala Lys Glu Leu Gly
465 470 475 480
Asn Gly Cys Phe Glu Phe Tyr His Lys Cys Asp Asn Glu Cys Met Glu
485 490 495
Ser Val Lys Asn Gly Thr Tyr Asp Tyr Pro Gln Tyr Ser Glu Glu Ala
500 505 510
Arg Leu Asn Arg Glu Glu Ile Ser Gly Val Lys Leu Glu Ser Met Gly
515 520 525
Thr Tyy Gln Ile Leu Ser Ile Tyr Ser Thr Val Ala Ser Ser Leu Ala
530 535 540
Leu Ala Ile Met Val Ala Gly Leu Ser Leu Trp Met Cys Ser Asn Gly
545 550 555 560
Ser Leu Gln Cys Arg Ile Cys Ile
565
.
<210>3
<211>101
<212>DNA
<213>人工合成
<400>3
agcgaaagca ggggtccaat ctgtcaaaat ggagaagatc gtgctgctgc tggccatcgt 60
gagcctggtg aagagcgacc agatctgcat cggctaccac g 101
<210>4
<211>90
<212>DNA
<213>人工合成
<400>4
I040426-序列表
ccagatctgc atcggctacc acgccaacaa cagcaccgag caggtggaca ccatcatgga 60
gaagaacgtg accgtgaccc acgcccagga 90
<210>5
<211>101
<212>DNA
<213>人工合成
<400>5
gccacgctgc agtcgcgcag gatcaggggc ttcacgccgt tcaggtcgca cagcttgccg 60
ttgtgggtct tctccaggat gtcctgggcg tgggtcacgg t 101
<210>6
<211>110
<212>DNA
<213>人工合成
<400>6
tgcagcgtgg ccggctggct gctgggcaac cccatgtgcg acgagttcat caacgtgccc 60
gagtggagct acatcgtgga gaaggccagc cccgccaacg acctgtgcta 110
<210>7
<211>110
<212>DNA
<213>人工合成
<400>7
ctgctcttgg ggatgatctg gatcttctcg aagtggttgg tgcggctcag caggtgcttc 60
agctcctcgt agtcgttgaa gtcgccgggg tagcacaggt cgttggcggg 110
<210>8
<211>98
<212>DNA
<213>人工合成
<400>8
caagagcagc tggagcaacc acgacgccag cagcggcgtg agcagcgcct gcccctacca 60
cggccgcagc agcttcttcc gcaacgtggt gtggctga 98
<210>9
<211>100
<212>DNA
I040426-序列表
<213>人工合成
<400>9
tgccccacag caccagcagg tcctcctggt tggtgttgtt gtagctgcgc ttgatggtgg 60
ggtaggcgct gttcttcttg atcagccaca ccacgttgcg 100
<210>10
<211>97
<212>DNA
<213>人工合成
<400>10
aggaggacct gctggtgctg tggggcatcc accaccccaa cgacgccgcc gagcagacca 60
agctgtacca gaaccccacc acctacatca gcgtggg 97
<210>11
<211>99
<212>DNA
<213>人工合成
<400>11
ctccatgcgg ccgctctgcc cgttcacctt ggggcgggtg gcgatctcgg gcaccaggcg 60
ctggttcagg gtgctggtgc ccacgctgat gtaggtggt 99
<210>12
<211>110
<212>DNA
<213>人工合成
<400>12
gcggccgcat ggagttcttc tggaccatcc tgaagcccaa cgacgccatc aacttcgaga 60
gcaacggcaa cttcatcgcc cccgagtacg cctacaagat cgtgaagaag 110
<210>13
<211>110
<212>DNA
<213>人工合成
<400>13
catgctgctg ttgatggcgc ccatgggggt ctggcacttg gtgttgcagt tgccgtactc 60
cagctcgctc ttcatgatgg cgctgtcgcc cttcttcacg atcttgtagg 110
<210>14
I040426-序列表
<211>110
<212>DNA
<213>人工合成
<400>14
gcatgccctt ccacaacatc caccccctga ccatcggcga gtgccccaag tacgtgaaga 60
gcaaccgcct ggtgctggcc accggcctgc gcaacacccc ccagcgcgag 110
<210>15
<211>110
<212>DNA
<213>人工合成
<400>15
gtacccgtac cagccgtcca ccatgccctg ccagccgccc tcgatgaagc cggcgatggc 60
gccgaacagg ccgcgcttct tgcggcggcg ctcgcgctgg ggggtgttgc 110
<210>16
<211>91
<212>DNA
<213>人工合成
<400>16
ggtacgggta ccaccacagc aacgagcagg gcagcggcta cgccgccgac aaggagagca 60
cccagaaggc catcgacggc gtgaccaaca a 91
<210>17
<211>103
<212>DNA
<213>人工合成
<400>17
gcggcggctc ttcctccagg ttgttgaact cgcggcccac ggcctcgaac tgggtgttca 60
tcttgtcgat gatgctgttc accttgttgg tcacgccgtc gat 103
<210>18
<211>97
<212>DNA
<213>人工合成
<400>18
caacctgctc ttcggagcgc cgcatcgaga acctgaacaa gcagatggag gacggcttcc 60
I040426-序列表
tggacgtgtg gacctacaac gccgagctgc tggtgct 97
<210>19
<211>99
<212>DNA
<213>人工合成
<400>19
gcgcagctgc aggcgcacct tgtcgtacag gttcttcacg ttgctgtcgt ggaagtccag 60
ggtgcgctcg ttctccatca gcaccagcag ctcggcgtt 99
<210>20
<211>99
<212>DNA
<213>人工合成
<400>20
gtgcgcctgc agctgcgcga caacgccaag gagctgggca acggctgctt cgagttctac 60
cacaagtgcg acaacgagtg catggagagc gtgaagaac 99
<210>21
<211>99
<212>DNA
<213>人工合成
<400>21
tgctctcgag cttcacgccg ctgatctcct cgcggttcag gcgggcctcc tcgctgtact 60
gggggtagtc gtaggtgccg ttcttcacgc tctccatgc 99
<210>22
<211>107
<212>DNA
<213>人工合成
<400>22
gtgaagctcg agagcatggg cacctaccag atcctgagca tctacagcac cgtggccagc 60
agcctggccc tggccatcat ggtggccggc ctgagcctgt ggatgtg 107
<210>23
<211>101
<212>DNA
<213>人工合成
I040426-序列表
<400>23
agtagacaca agggtgtttt taactacaat ctgaactcac aaatttagat gcagatgcgg 60
cactgcaggc tgccgttgct gcacatccac aggctcaggc c 101
<210>24
<211>26
<212>DNA
<213>人工合成
<400>24
agcgaaagca ggggtccaat ctgtca 26
<210>25
<211>26
<212>DNA
<213>人工合成
<400>25
ccagatctgc atcggctacc acgcca 26
<210>26
<211>25
<212>DNA
<213>人工合成
<400>26
gccacgctgc agtcgcgcag gatca 25
<210>27
<211>27
<212>DNA
<213>人工合成
<400>27
gactgcagcg tggccggctg gctgctg 27
<210>28
<211>28
<212>DNA
<213>人工合成
I040426-序列表
<400>28
ccagctgctc ttggggatga tctggatc 28
<210>29
<211>24
<212>DNA
<213>人工合成
<400>29
caagagcagc tggagcaacc acga 24
<210>30
<211>25
<212>DNA
<213>人工合成
<400>30
tgccccacag caccagcagg tcctc 25
<210>31
<211>23
<212>DNA
<213>人工合成
<400>31
aggaggacct gctggtgctg tgg 23
<210>32
<211>25
<212>DNA
<213>人工合成
<400>32
ctccatgcgg ccgctctgcc cgttc 25
<210>33
<211>27
<212>DNA
<213>人工合成
<400>33
gagcggccgc atggagttct tctggac 27
<210>34
I040426-序列表
<211>27
<212>DNA
<213>人工合成
<400>34
ggcatgctgc tgttgatggc gcccatg 27
<210>35
<211>25
<212>DNA
<213>人工合成
<400>35
cagcatgccc ttccacaaca tccac 25
<210>36
<211>26
<212>DNA
<213>人工合成
<400>36
gtggtacccg taccagccgt ccacca 26
<210>37
<211>25
<212>DNA
<213>人工合成
<400>37
ggtacgggta ccaccacagc aacga 25
<210>38
<211>24
<212>DNA
<213>人工合成
<400>38
gcggcggctc ttcctccagg ttgt 24
<210>39
<211>25
<212>DNA
I040426-序列表
<213>人工合成
<400>39
caacctgctc ttcggagcgc cgcat 25
<210>40
<211>24
<212>DNA
<213>人工合成
<400>40
gcgcagctgc aggcgcacct tgtc 24
<210>41
<211>24
<212>DNA
<213>人工合成
<400>41
gtgcgcctgc agctgcgcga caac 24
<210>42
<211>23
<212>DNA
<213>人工合成
<400>42
tgctctcgag cttcacgccg ctg 23
<210>43
<211>23
<212>DNA
<213>人工合成
<400>43
gtgaagctcg agagcatggg cac 23
<210>44
<211>27
<212>DNA
<213>人工合成
<400>44
I040426-序列表
agtagacaca agggtgtttt taactac 27
<210>45
<211>21
<212>DNA
<213>人工合成
<400>45
attaaccctc actaaaggga a 21
<210>46
<211>20
<212>DNA
<213>人工合成
<400>46
acttaatacg actcactata 20
<210>47
<211>26
<212>DNA
<213>人工合成
<400>47
ccactctaga gtgtttttaa ctacaa 26
<210>48
<211>20
<212>DNA
<213>人工合成
<400>48
acttaatacg actcactata 20
<210>49
<211>18
<212>DNA
<213>人工合成
<400>49
gcaccaacca cttcgaga 18
<210>50
I040426-序列表
<211>20
<212>DNA
<213>人工合成
<400>50
gctaccacgc caacaacagc 20
<210>51
<211>19
<212>DNA
<213>人工合成
<400>51
gaagagcgag ctggagtac 19
<210>52
<211>21
<212>DNA
<213>人工合成
<400>52
gcagatggag gacggcttcc t 21
<210>53
<211>20
<212>DNA
<213>人工合成
<400>53
tgcagcttat aatggttaca 20
<210>54
<211>22
<212>DNA
<213>人工合成
<400>54
tcaaaatgga gaagatcgtg ct 22
<210>55
<211>26
<212>DNA
<213>人工合成
I040426-序列表
<400>55
ccactctaga gtgtttttaa ctacaa 26
Claims (9)
1.一种禽流感病毒血凝素基因，其特征在于具有SEQ ID NO：1所示的核苷酸序列。
2.一种表达载体，其特征在于包含权利要求1所述的基因。
3.权利要求2所述的表达载体，其是pCIoptiHA。
4.一种宿主细胞，其特征在于表达权利要求2-3任一项所述的表达载体。
5.权利要求4所述的宿主细胞，其是真核细胞。
6.权利要求5所述的宿主细胞，其是人胚胎肾细胞、COS细胞、鸡胚成纤维细胞。
7.权利要求1所述的基因在制备H5亚型流感病毒DNA疫苗中作为免疫原基因的应用。
8.权利要求1所述的基因在制备亚单位疫苗、重组禽痘病毒基因工程疫苗中作为免疫原基因的应用。
9.权利要求1所述的基因在制备单克隆抗体中作为免疫原基因的应用。
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CNB2004100098665A CN1326998C (zh) | 2004-11-25 | 2004-11-25 | 编码h5亚型禽流感病毒血凝素蛋白的基因及其应用 |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CNB2004100098665A CN1326998C (zh) | 2004-11-25 | 2004-11-25 | 编码h5亚型禽流感病毒血凝素蛋白的基因及其应用 |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| CN1632124A true CN1632124A (zh) | 2005-06-29 |
| CN1326998C CN1326998C (zh) | 2007-07-18 |
Family
ID=34845530
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CNB2004100098665A Expired - Lifetime CN1326998C (zh) | 2004-11-25 | 2004-11-25 | 编码h5亚型禽流感病毒血凝素蛋白的基因及其应用 |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN1326998C (zh) |
Cited By (16)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101460628A (zh) * | 2006-05-05 | 2009-06-17 | 淡马锡生命科学研究院有限公司 | 生物分子表面展示及其用途 |
| CN101070347B (zh) * | 2006-05-12 | 2010-06-30 | 青岛宝麦德生物医药科技有限公司 | 禽流感h5n1新型粘膜免疫疫苗及其应用 |
| CN101693894B (zh) * | 2009-10-15 | 2011-07-27 | 北京大学深圳研究生院 | 禽流感病毒h5n1假病毒及其制备方法与专用dna片段 |
| CN101186637B (zh) * | 2007-11-14 | 2011-09-14 | 中国科学院微生物研究所 | 抑制流感病毒感染的方法及其药物 |
| CN102603874A (zh) * | 2012-03-28 | 2012-07-25 | 广西壮族自治区兽医研究所 | 用于检测h5亚型禽流感病毒的重组ha蛋白及其编码基因 |
| CN102645539A (zh) * | 2012-03-28 | 2012-08-22 | 广西壮族自治区兽医研究所 | H5亚型禽流感病毒抗体的ha抗原表位-elisa试剂盒 |
| CN102645538A (zh) * | 2012-03-28 | 2012-08-22 | 广西壮族自治区兽医研究所 | H5亚型禽流感病毒抗体的ha抗原表位胶体金快速检测试纸条 |
| CN102973952A (zh) * | 2012-11-28 | 2013-03-20 | 中国农业科学院哈尔滨兽医研究所 | 表达传染性法氏囊病病毒多聚蛋白基因vp243的dna疫苗及其构建方法和应用 |
| CN103169963A (zh) * | 2013-04-12 | 2013-06-26 | 中国农业科学院哈尔滨兽医研究所 | 狂犬病dna疫苗的构建及应用 |
| CN103421843A (zh) * | 2013-07-29 | 2013-12-04 | 中国农业科学院哈尔滨兽医研究所 | 编码h5n1亚型禽流感同义血凝素(ha)蛋白以及同义神经氨酸酶(na)蛋白的基因及其应用 |
| CN101851289B (zh) * | 2008-10-09 | 2015-01-07 | 厦门大学 | H5亚型禽流感病毒血凝素蛋白的人源化抗体及其用途 |
| CN106039303A (zh) * | 2016-06-20 | 2016-10-26 | 华南农业大学 | 一种h5亚型禽流感dna疫苗及其制备方法 |
| CN109439633A (zh) * | 2018-11-09 | 2019-03-08 | 山东信得科技股份有限公司 | 一种插入h7n9的ha蛋白的新城疫病毒重组疫苗株 |
| CN109439687A (zh) * | 2018-11-09 | 2019-03-08 | 山东信得科技股份有限公司 | 一种表达禽流感h9n2病毒ha蛋白的新城疫病毒载体疫苗株 |
| CN111526886A (zh) * | 2017-10-27 | 2020-08-11 | 国家血清研究所 | 多基因流感疫苗 |
| JP2022512881A (ja) * | 2018-11-06 | 2022-02-07 | ベーリンガー インゲルハイム フェトメディカ ゲーエムベーハー | 鳥インフルエンザウイルスｈ５亜型に対する免疫原性組成物 |
Family Cites Families (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CA2524293A1 (en) * | 2003-05-05 | 2004-11-18 | Boyce Thompson Institute For Plant Research | Vectors and cells for preparing immunoprotective compositions derived from transgenic plants |
| CN1263848C (zh) * | 2003-12-02 | 2006-07-12 | 中国农业科学院哈尔滨兽医研究所 | 人工重组的h9n2亚型流感病毒及其应用 |
-
2004
- 2004-11-25 CN CNB2004100098665A patent/CN1326998C/zh not_active Expired - Lifetime
Cited By (26)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101460628B (zh) * | 2006-05-05 | 2013-03-27 | 淡马锡生命科学研究院有限公司 | 生物分子表面展示及其用途 |
| CN101460628A (zh) * | 2006-05-05 | 2009-06-17 | 淡马锡生命科学研究院有限公司 | 生物分子表面展示及其用途 |
| CN101070347B (zh) * | 2006-05-12 | 2010-06-30 | 青岛宝麦德生物医药科技有限公司 | 禽流感h5n1新型粘膜免疫疫苗及其应用 |
| CN101186637B (zh) * | 2007-11-14 | 2011-09-14 | 中国科学院微生物研究所 | 抑制流感病毒感染的方法及其药物 |
| CN101851289B (zh) * | 2008-10-09 | 2015-01-07 | 厦门大学 | H5亚型禽流感病毒血凝素蛋白的人源化抗体及其用途 |
| CN101693894B (zh) * | 2009-10-15 | 2011-07-27 | 北京大学深圳研究生院 | 禽流感病毒h5n1假病毒及其制备方法与专用dna片段 |
| CN102645538B (zh) * | 2012-03-28 | 2014-05-07 | 广西壮族自治区兽医研究所 | H5亚型禽流感病毒抗体的ha抗原表位胶体金快速检测试纸条 |
| CN102603874A (zh) * | 2012-03-28 | 2012-07-25 | 广西壮族自治区兽医研究所 | 用于检测h5亚型禽流感病毒的重组ha蛋白及其编码基因 |
| CN102645538A (zh) * | 2012-03-28 | 2012-08-22 | 广西壮族自治区兽医研究所 | H5亚型禽流感病毒抗体的ha抗原表位胶体金快速检测试纸条 |
| CN102603874B (zh) * | 2012-03-28 | 2014-07-02 | 广西壮族自治区兽医研究所 | 用于检测h5亚型禽流感病毒的重组ha蛋白及其编码基因 |
| CN102645539A (zh) * | 2012-03-28 | 2012-08-22 | 广西壮族自治区兽医研究所 | H5亚型禽流感病毒抗体的ha抗原表位-elisa试剂盒 |
| CN102645539B (zh) * | 2012-03-28 | 2014-04-16 | 广西壮族自治区兽医研究所 | H5亚型禽流感病毒抗体的ha抗原表位-elisa试剂盒 |
| CN102973952A (zh) * | 2012-11-28 | 2013-03-20 | 中国农业科学院哈尔滨兽医研究所 | 表达传染性法氏囊病病毒多聚蛋白基因vp243的dna疫苗及其构建方法和应用 |
| CN102973952B (zh) * | 2012-11-28 | 2015-02-04 | 中国农业科学院哈尔滨兽医研究所 | 表达传染性法氏囊病病毒多聚蛋白基因vp243的dna疫苗及其构建方法和应用 |
| CN103169963A (zh) * | 2013-04-12 | 2013-06-26 | 中国农业科学院哈尔滨兽医研究所 | 狂犬病dna疫苗的构建及应用 |
| CN103169963B (zh) * | 2013-04-12 | 2014-10-15 | 中国农业科学院哈尔滨兽医研究所 | 狂犬病dna疫苗的构建及应用 |
| CN103421843A (zh) * | 2013-07-29 | 2013-12-04 | 中国农业科学院哈尔滨兽医研究所 | 编码h5n1亚型禽流感同义血凝素(ha)蛋白以及同义神经氨酸酶(na)蛋白的基因及其应用 |
| CN103421843B (zh) * | 2013-07-29 | 2015-12-09 | 中国农业科学院哈尔滨兽医研究所 | 编码h5n1亚型禽流感同义血凝素(ha)蛋白以及同义神经氨酸酶(na)蛋白的基因及其应用 |
| CN106039303A (zh) * | 2016-06-20 | 2016-10-26 | 华南农业大学 | 一种h5亚型禽流感dna疫苗及其制备方法 |
| CN111526886A (zh) * | 2017-10-27 | 2020-08-11 | 国家血清研究所 | 多基因流感疫苗 |
| JP2022512881A (ja) * | 2018-11-06 | 2022-02-07 | ベーリンガー インゲルハイム フェトメディカ ゲーエムベーハー | 鳥インフルエンザウイルスｈ５亜型に対する免疫原性組成物 |
| JP7337921B2 (ja) | 2018-11-06 | 2023-09-04 | ベーリンガー インゲルハイム フェトメディカ ゲーエムベーハー | 鳥インフルエンザウイルスｈ５亜型に対する免疫原性組成物 |
| CN109439633A (zh) * | 2018-11-09 | 2019-03-08 | 山东信得科技股份有限公司 | 一种插入h7n9的ha蛋白的新城疫病毒重组疫苗株 |
| CN109439687A (zh) * | 2018-11-09 | 2019-03-08 | 山东信得科技股份有限公司 | 一种表达禽流感h9n2病毒ha蛋白的新城疫病毒载体疫苗株 |
| CN109439687B (zh) * | 2018-11-09 | 2022-01-04 | 山东信得科技股份有限公司 | 一种表达禽流感h9n2病毒ha蛋白的新城疫病毒载体疫苗株 |
| CN109439633B (zh) * | 2018-11-09 | 2022-01-07 | 山东信得科技股份有限公司 | 一种插入h7n9的ha蛋白的新城疫病毒重组疫苗株 |
Also Published As
| Publication number | Publication date |
|---|---|
| CN1326998C (zh) | 2007-07-18 |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| CN1282656C (zh) | 来自gagp17和p24保守区域的HIV肽以及它们在例如疫苗中的应用 | |
| CN1188519C (zh) | 包含HIV－1Tat和/或Nef蛋白的融合蛋白 | |
| CN1298738C (zh) | 修饰的麻疹病毒v蛋白 | |
| CN1632124A (zh) | 编码h5亚型禽流感病毒血凝素蛋白的基因及其应用 | |
| CN1602316A (zh) | 黄病毒ns1亚单位疫苗 | |
| CN1720060A (zh) | 人乳头瘤病毒多肽和免疫原性组合物 | |
| CN1346367A (zh) | Hiv肽,抗原,疫苗组合物,检测hiv所诱导的抗体的免疫测定试剂盒及方法 | |
| CN1829799A (zh) | 含polii启动子和核酶的重组流感载体 | |
| CN1556857A (zh) | 呼吸道合胞病毒(rsv)g蛋白的新肽及其在疫苗中的应用 | |
| CN101054582A (zh) | A型肉毒毒素受体结合区Hc基因及其编码蛋白与应用 | |
| CN1069068A (zh) | 重组禽痘病毒，培养被上述病毒感染的细胞以及从上述病毒得到的家禽疫苗 | |
| CN1458936A (zh) | 来自Tat、Rev和Nef保守区的HIV肽以及它们作为例如疫苗组分的应用 | |
| CN1990869A (zh) | 鸡传染性法氏囊病病毒VP2 cDNA、其表达载体、所表达的重组蛋白及应用 | |
| CN1898390A (zh) | 用于在改良安卡拉痘苗病毒中进行表达的启动子 | |
| CN1718243A (zh) | 一类免疫佐剂及其在抗病毒疫苗或药物制备中的应用 | |
| CN1618464A (zh) | 用于抗日本脑炎病毒感染的灭活疫苗的增强免疫原及其生产方法 | |
| CN1314806C (zh) | 小型化抗eb病毒肿瘤多肽及其应用与制备方法 | |
| CN1273593C (zh) | 丙型肝炎病毒多表位抗原的基因克隆及其编码序列 | |
| CN1531548A (zh) | 编码可诱导抗病毒效应的蛋白质的嵌合链 | |
| CN1922309A (zh) | 具有反式互补的基因组缺陷的呼吸道合胞病毒 | |
| CN1244698C (zh) | 一种重组sars病毒基因在多形汉逊酵母中的表达及用途 | |
| CN100339396C (zh) | 抑制i型人免疫缺陷病毒感染的三螺旋蛋白及其编码基因与应用 | |
| CN1517437A (zh) | 特异性治疗肿瘤或胞内感染的疫苗及应用 | |
| CN101039955A (zh) | 与sars冠状病毒刺突蛋白相关的核酸、多肽、表达方法和免疫原性组合物 | |
| CN1163604C (zh) | 减毒水痘病毒冈株的基因62和利用基因62的减毒水痘活疫苗用病毒株的鉴定方法 |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| C06 | Publication | ||
| PB01 | Publication | ||
| C10 | Entry into substantive examination | ||
| SE01 | Entry into force of request for substantive examination | ||
| C14 | Grant of patent or utility model | ||
| GR01 | Patent grant | ||
| EE01 | Entry into force of recordation of patent licensing contract |
Application publication date: 20050629 Assignee: HARBIN WEIKE BIOTECHNOLOGY DEVELOPMENT Co.,Ltd. Assignor: HARBIN VETERINARY Research Institute CHINESE ACADEMY OF AGRICULTURE SCIENCE Contract record no.: 2014230000506 Denomination of invention: Gene encoding hemagglutinin protein of H5 avian influenza virus and its application Granted publication date: 20070718 License type: Exclusive License Record date: 20140911 |
|
| LICC | Enforcement, change and cancellation of record of contracts on the licence for exploitation of a patent or utility model | ||
| CX01 | Expiry of patent term | ||
| CX01 | Expiry of patent term |
Granted publication date: 20070718 |