CN106011086A - Expression-signal-peptide-substituted recombinant Newcastle disease heat-resistant vaccine strain of respiratory syncytial virus F protein and preparation method - Google Patents
Expression-signal-peptide-substituted recombinant Newcastle disease heat-resistant vaccine strain of respiratory syncytial virus F protein and preparation method Download PDFInfo
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- C12N2760/18011—Paramyxoviridae
- C12N2760/18511—Pneumovirus, e.g. human respiratory syncytial virus
- C12N2760/18534—Use of virus or viral component as vaccine, e.g. live-attenuated or inactivated virus, VLP, viral protein
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- C12N2800/10—Plasmid DNA
- C12N2800/106—Plasmid DNA for vertebrates
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Abstract
The invention discloses an expression-signal-peptide-substituted recombinant Newcastle disease heat-resistant vaccine strain of respiratory syncytial virus F protein and a preparation method. The classified name of the heat-resistant vaccine strain is Rts-tF/RSV, and the heat-resistant vaccine strain is preserved in China Center For Type Culture Collection in Wuhan University on April 19th, 2016 with the preservation number being CCTCC NO: V201625. The vaccine strain can efficiently express truncated F protein of the respiratory syncytial virus after being infected with animals or cells, and tPAs carried with the vaccine strain can enhance secretory expression and immunogenicity of the truncated F protein. The strain can be used for preparing respiratory syncytial virus sigmasubunit heat-resistant live vaccines.
Description
Technical field
The invention belongs to biology field, be specifically related to the respiratory syncystial virus F that a kind of expression signal peptide is replaced
The heat-resisting vaccine strain of recombinant Newcastle disease of albumen and preparation method.
Background technology
Respiratory syncytial virus (Respiratory syncytial virus, RSV) is to cause infant lower respiratory tract sense
The important pathogen of dye, causes bronchitis and pneumonia, can cause death time serious, to the health and lives seat belt of infant
Carry out grave danger.Vaccination is the effective means of prophylaxis of viral diseases.In order to effectively prevent rsv infection disease, people
Always at RSV vaccine development, but secure permission listing still without vaccine at present.Therefore, researching and developing safely and effectively RSV vaccine is
The top priority of prevention rsv infection.Be positioned at RSV virion surface glycoprotein fusion protein (fusion protein,
F) it is its main protective antigen, is also one of first-selected component developing RSV subunit vaccine.
Along with the continuous progress of Protocols in Molecular Biology, the research and development of novel gene engineered vaccine constantly make a breakthrough, wherein
Novel multi-connected live vaccine based on Newcastle Disease poisonous carrier is one of research and development focus, and the immunogenic gene of many cause of diseases exists
Successful expression in newcastle carrier, and achieve preferable immune protective effect, as infectious bursa of Fabricius virus VP2 albumen,
The HA albumen of H5 subtype avian influenza virus, the G glycoprotein of rabies virus, the CoV albumen of SARS virus, people
The Gag albumen etc. of para-immunity defective virus.This type of vaccine has inducing systemic immunity (humoral immunization, cellular immunization and viscous
Film immunity), high Embryo Gallus domesticus growth characteristics, low production cost, immunization ways easy modes such as () drinking-water, aerosol, eye dripping/collunariums,
The advantages such as safety (occurring restructuring and virulence to return strong probability between strain minimum).But existing Avian pneumo-encephalitis virus carrier bacterin is all
With nonrefractory type Avian pneumo-encephalitis virus as framework construction.
The immunogenicity of vaccine relies primarily on antigen levels and antigen presentation efficiency.For in theory, it is positioned Cytoplasm
Antigen can more effectively induce immunoreation.Tissue-typed plasminogen activator signal sequence (tPAs) be one special
Albumen sorting signals, it directly sorts the antigen expressed to endoplasmic reticulum.After merging with tPAs, Japanese B encephalitis virus and cattle
Expression in the envelope protein cell after transfection of viral diarrhea disease virus is significantly higher than wild-type protein, and mainly
It is positioned on Cytoplasm and cell membrane, higher humoral immunization and cellular immunization can be caused.
In view of the RSV high hazardness to infant, people have put into substantial amounts of energy and have carried out RSV vaccine development.But due to
The high-risk group that this virus infects is the most jejune baby of developing immune system, adds and vaccine enhancement disease easily occurs,
So it is difficult to research and develop this viral vaccine.Even so, people have also been an attempt to kinds of schemes, including attenuated live epidemic disease
Seedling, subunit vaccine, vector-viral vaccine, virus-like particle (Virus Like Particle, VLP) vaccine, DNA vaccination etc..
Therefore, how to provide a kind of have that the expression signal peptide that heat-resisting, stability is strong, refrigerated condition requirement is low replaces exhale
The heat-resisting vaccine strain of recombinant Newcastle disease of suction road syncystial virus F protein and preparation method are the skills that those skilled in the art are urgently to be resolved hurrily
Art problem.
Summary of the invention
For deficiency of the prior art, the present invention conducts in-depth research, it is provided that a kind of expression signal peptide is replaced
The heat-resisting vaccine strain of recombinant Newcastle disease of respiratory syncystial virus F protein and preparation method.
The recombinant Newcastle disease that one aspect of the present invention relates to the respiratory syncystial virus F protein that a kind of expression signal peptide is replaced is resistance to
Hot vaccine strain, it is characterised in that described heat-resisting vaccine strain Classification And Nomenclature is rTS-tF/RSV, was preserved on April 19th, 2016
Wuhan University's China typical culture collection center, preserving number is CCTCC NO:V201625.
The recombinant Newcastle disease that another aspect of the present invention further relates to the respiratory syncystial virus F protein that expression signal peptide is replaced is resistance to
The preparation method of hot vaccine strain, it is characterised in that comprise the steps:
The amplification of the porcine circovirus 2 type Cap gene that 1.1 signal peptides are replaced
Being bred on Hep-2 cell by Respiratory Syncytial Virus Strain Long, after inoculating 5 days, multigelation results are thin
Born of the same parents' culture fluid.The cell culture fluid of results is carried out the PCR amplification of truncate F gene, it is thus achieved that the F protein of truncate；With primer from prolonging
The method amplification stretched obtains tPAs sequence；Truncate F gene order and tPAs sequence are attached by the way of over-lap PCR,
Carrying out PCR amplification again, amplimer is: forward primer 5 '-CAGCTATATTAAGGATTAAGAAAAAATACGGGTAGAAAGCT
TGCCACCATGGATGC-3 ', downstream primer 5 '-GGGGCACTCCGATTCTACCCGTATTTTTTCTTAATTATTATTAATTT
GTGGTTG-3′；PCR primer carrying out agarose gel reclaim after purification, measure DNA concentration, pending clone connects；
The PCR amplification of 1.2 Avian pneumo-encephalitis virus heat-resistant carriers fragments
Obtaining Avian pneumo-encephalitis virus carrier sequence with the method for super LA-PCR, amplimer is: forward primer 5 '-
GAATCGGAGTGCCCCGATTGTGCCAAGATGGACTCATC-3 ', downstream primer 5 '-TCCTTAATATAGCTGAATTGATTG
CAGCTGCGCGATC-3′；Amplification template is Avian pneumo-encephalitis virus heat-resisting strain plasmid pTS09-C；Carry out PCR primer reclaiming purification,
Measure its concentration, treat that clone connects；
The clone of 1.3 recombiant plasmid pTS-tF/RSV connects
Use In-fusion to clone interconnection technique, truncate F genetic fragment good for purification and heat-resistant carriers fragment are carried out gram
Grand connection (catenation sequence is CAGCTATATTAAGGA and TCGGAGTGCCCCGAT)；Connect product conversion DH5 α competence thin
Born of the same parents, the cell after conversion is chosen single bacterium colony and is carried out liquid culture, is accredited as positive bacterium solution and enters after cultivating 16 hours on LB plate
Row plasmid extraction；
1.4 recombinant virus rTS-tF/RSV rescues recover
Use lipofection, the recombiant plasmid pTS-tF/RSV built and three are expressed NP, P and L egg respectively
White helper plasmid cotransfection BHK-21 cell；Transfect latter 72 hours, multigelation harvesting liquid, inoculate 9-11 age in days SPF
Embryo Gallus domesticus.Inoculate latter 5 days, gather in the crops chick embryo allantoic liquid, the weight of the respiratory syncystial virus F protein that isolated expression signal peptide is replaced
The group heat-resisting vaccine strain of newcastle.
The invention still further relates to the resistance to epidemic disease due to heat pathogen of recombinant Newcastle disease of the respiratory syncystial virus F protein that above-mentioned expression signal peptide is replaced
Seedling strain application in preparing recombinant virus.
The invention still further relates to the resistance to epidemic disease due to heat pathogen of recombinant Newcastle disease of the respiratory syncystial virus F protein that above-mentioned expression signal peptide is replaced
Seedling strain application in preparing the heat-resisting live vaccine of respiratory syncytial virus sub-units.
This application discloses the resistance to epidemic disease due to heat pathogen of recombinant Newcastle disease of the respiratory syncystial virus F protein that a kind of expression signal peptide is replaced
Miao Zhu, this vaccine strain has significant heat-resistant quality, can greatly reduce the dependence to cold chain system, save preserve cost of transportation and
Improve vaccine heat stability.This strain, can the truncate F egg of high efficient expression respiratory syncytial virus after infection animal or cell
In vain, the tPAs carried can strengthen secreting, expressing and the immunogenicity of truncate F protein.It is sick that this strain can be used for preparing respiratory syncystial
The poison heat-resisting live vaccine of subunit.
Accompanying drawing explanation
Fig. 1 is the recombinant heat-proof Newcastle Disease of the respiratory syncytial virus truncate F protein that expression signal peptide of the present invention is replaced
The structure schematic diagram of poison total length plasmid.Wherein rTS09-C is the genome structure of parent plant, and rTS-tF/RSV is at rTS09-C
Genome inserts the structure after the truncate F protein that signal peptide is replaced, is labelled with the overall sequence composition of insertion sequence in detail.
Detailed description of the invention
Below in conjunction with embodiment, the present invention is described further, the following stated, is only the preferable enforcement to the present invention
Example, not does the restriction of other forms to the present invention, and any those skilled in the art are possibly also with the disclosure above
Technology contents be changed to the Equivalent embodiments that changes on an equal basis.Every without departing from the present invention program content, according to the present invention
Technical spirit any simple modification that following example are done or equivalent variations, the most within the scope of the present invention.
Embodiment 1
The first step, the structure of recombinant full-lenght plasmid pTS-tF/RSV and qualification
The amplification of the porcine circovirus 2 type Cap gene that 1.1 signal peptides are replaced
Being bred on Hep-2 cell by Respiratory Syncytial Virus Strain Long, after inoculating 5 days, multigelation results are thin
Born of the same parents' culture fluid.The cell culture fluid of results is carried out the PCR amplification of truncate F gene, it is thus achieved that F protein (the erasure signal peptide of truncate
With cross-film district).TPAs sequence is obtained from the method amplification extended with primer.Truncate F gene order and tPAs sequence are passed through weight
The mode of folded PCR is attached, then carries out PCR amplification, and amplimer is: forward primer 5 '-CAGCTATATTAAGGATTAAGA
AAAAATACGGGTAGAAAGCTTGCCACCATGGATGC-3 ', downstream primer 5 '-GGGGCACTCCGATTCTACCCGTATTTT
TTCTTAATTATTATTAATTTGTGGTTG-3 ' (underscore part is the complementary series connected for In-fusion clone).
Agarose gel electrophoresis testing goal band about 1.7kb, is consistent with intended 1681bp and (deletes native signal peptide and cross-film district
F mrna length is 1512bp, introduces gene start sequence and gene termination sequence in F gene front-end and back-end respectively, simultaneously
Kozak sequence and tPAs sequence is introduced) in F gene front end.PCR primer carries out agarose gel reclaim after purification, measure
DNA concentration, pending clone connects.
The PCR amplification of 1.2 Avian pneumo-encephalitis virus heat-resistant carriers fragments
Obtaining Avian pneumo-encephalitis virus carrier sequence with the method for super LA-PCR, amplimer is: forward primer 5 '-
GAATCGGAGTGCCCCGATTGTGCCAAGATGGACTCATC-3 ', downstream primer 5 '-TCCTTAATATAGCTGAATTGATTG
CAGCTGCGCGATC-3 ' (underscore part is the complementary series connected for In-fusion clone).Amplification template is new city
The epidemic disease heat-resisting strain plasmid pTS09-C of poison.Agarose gel electrophoresis testing goal band about 18kb, with intended 17.8kb size phase
Symbol.Carry out PCR primer reclaiming purification, measure its concentration, treat that clone connects.
The clone of 1.3 recombiant plasmid pTS-tF/RSV connects
According to the mode shown in Fig. 1, use In-fusion to clone interconnection technique, by truncate F genetic fragment good for purification and
Heat-resistant carriers fragment carries out clone and connects.Connecting product and convert DH5 α competent cell, the cell after conversion is trained on LB plate
Choose single bacterium colony after supporting 16 hours and carry out liquid culture.The PCR that bacterium solution carries out truncate F gene identifies.It is accredited as the bacterium solution of the positive
Carry out plasmid extraction, treat that enzyme action and order-checking are identified.
The enzyme action of 1.4 recombiant plasmid pTS-tF/RSV is identified with order-checking
With BamHI and MluI restriction endonuclease, total length recombiant plasmid is carried out enzyme action qualification respectively, all obtained being consistent with expection
Restriction enzyme mapping.Enzyme action is identified, and correct recombiant plasmid pTS-tF/RSV delivers to order-checking company and carries out sequencing, result table
The truncate F gene of bright respiratory syncytial virus is inserted between P and the M gene of Avian pneumo-encephalitis virus heat-resistant carriers, tPAs sequence
Successfully substituted for the signal peptide sequence of F protein self, and all sequences is completely the same with expection, recombiant plasmid pTS-tF/RSV structure
Build up merit.
Second step, recombinant virus rTS-tF/RSV rescue recovers and identifies
The rescue of 2.1 recombinant virus rTS-tF/RSV recovers
Use lipofection, the recombiant plasmid pTS-tF/RSV built and three are expressed NP, P and L egg respectively
White helper plasmid cotransfection BHK-21 cell (infecting the vaccinia virus expressing t7 rna polymerase in advance).Transfect latter 72 hours,
Multigelation harvesting liquid, inoculates 9-11 age in days SPF Embryo Gallus domesticus.Inoculate latter 5 days, gather in the crops chick embryo allantoic liquid, pending newcastle
Viral diagnosis.
The hemagglutinative titer of 2.2 recombinant viruses and fluorescence quantitative PCR detection
With hemagglutination test and the method for quantitative fluorescent PCR, detecting results chick embryo allantoic liquid, result is newcastle
Positive.Tentatively show that recombinant virus rTS-tF/RSV saves successfully.
PCR amplification and the order-checking of 2.3 recombinant virus tF/RSV genes
The specificity amplification primer of application respiratory syncytial virus truncate F gene, cuts recombinant virus rTS-tF/RSV
Short F gene carries out RT-PCR amplification, and pcr amplification product is carried out Sequence analysis, obtains 1681 long truncate F gene orders
(SEQ ID No.1), holds from 5 ' to 3 ', contains the gene end of the part non-coding area sequence (UTR) of P gene, P gene successively
Only sequence (GE), intermediate sequence (IG), the gene start sequence (GS) of truncate F gene, Kozak sequence, tPAs sequence, truncate F
Gene order (deleting native signal peptide and cross-film district), the dual termination codon of truncate F gene, the GE sequence of truncate F gene
Row, the GS sequence of IG sequence, M gene and the UTR sequence of M gene.Consistent with expected sequence 100%.
The respiratory syncytial virus truncate F protein that 3rd step, signal peptide are replaced table in recombinant virus rTS-tF/RSV
Reach checking
The expression of the truncate F protein that 3.1 indirect immunofluorescence detection signal peptides are replaced
Recombinant virus rTS-tF/RSV and rTS09-C (comparison) is infected BHK-21 cell with 0.01MOI respectively, after 24h
Carrying out indirect immunofluorescene assay, one resists for NDV positive chicken serum or RSV positive rabbit anteserum.Testing result shows, recombinant virus
It is intracellular that rTS-tF/RSV infects, and uses NDV positive chicken serum or RSV positive rabbit anteserum to resist as one and green all can be detected
Fluorescence, and the cell that rTS09-C strain is infected only detects fluorescence under NDV positive serum effect.
The expression of the truncate F protein that 3.2Western Blot method detection signal peptide is replaced
Recombinant virus rTS-tF/RSV and rTS09-C (comparison) is infected BHK-21 cell with 0.01MOI respectively, after 24h
Collect cell pyrolysis liquid, be one anti-to carry out Western Blot detection with porcine circovirus 2 type positive serum.Testing result table
Bright, an obvious purpose band occurs near 60KDa, close with intended 58.68KDa size.Show that signal peptide is replaced
Truncate F protein in the intracellular correct expression of recombinant virus infection.
4th step, the Identification of Biological Characteristics of recombinant virus rTS-tF/RSV
The multiplication characteristic of 4.1 recombinant viruses measures
For the growing multiplication situation of heavier papova rTS-tF/RSV strain Yu parent's rTS09-C strain, two strain virus are divided
Not with 100TCID50Inoculation BHK-21 cell, after inoculation, 24h, 48h, 72h, 96h take supernatant 500 μ l, measure virus TCID50, paint
The growth curve of system virus.Result shows, recombinant virus rTS-tF/RSV strain and parent's rTS09-C strain are in the life of BHK-21 cell
Long Similar Broken Line, viral titer is close, and about 107.0TCID50/ml。
The Pathogenicity of 4.2 recombinant viruses
Measure the recombinant virus median lethal time (MDT/MLD) to the minimal lethal dose of Embryo Gallus domesticus, result display difference
Dilution factor (10-1To 10-9) virus inoculation Embryo Gallus domesticus after 120h in all will not be lethal to Embryo Gallus domesticus, recombinant virus rTS-tF/RSV
MDT/MLD more than 120h, show that recombinant virus maintains the low toxicity characteristic of parent plant, the insertion of exogenous gene does not change weight
Papova pathogenic.
The heat-resistant quality of 4.3 recombinant viruses measures
Measure recombinant virus HA heat-resistant quality under 56 DEG C of environment, nonrefractory strain LaSota strain and heat-resisting strain are set simultaneously
RTS09-C strain is comparison.As shown in table 1, nonrefractory strain LaSota is after 56 DEG C of 9min of resistance to heat treatment, and HA titer reduces to 0, parent
Strain rTS09-C and recombinant virus rTS-tF/RSV still has hemagglutination activity after 56 DEG C of 150min of resistance to heat treatment.
Show that the heat-resistant quality of recombinant virus rTS-tF/RSV is consistent with parent plant, be heat-resisting strain.(this Strain is
Preservation, is deposited in Wuhan University's China typical culture collection center, and preserving number is CCTCC NO:V201625).
The HA heat-resistant quality measurement result of table 1. recombinant virus
aRepresent that virus has hemagglutination activity,bRepresent that virus does not has hemagglutination activity
Embodiment described above is the preferred embodiments of the present invention, and does not limit the practical range of the present invention.Therefore, all
It is according to the equivalent modifications done by the essence of present invention, all should belong within the scope of the present invention.
<110>Animal Husbandry and Veterinary Inst., Hubei Academy of Agricultural Sciences
<120>expression signal peptide is replaced the heat-resisting vaccine strain of recombinant Newcastle disease of respiratory syncystial virus F albumen and preparation method
<160>1
Sequence
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atctcttcat ttgcttctag tcaaaacatc actgaagaat tttatcaatc aacatgcagt 180
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ggttttttgt taggtgttgg atctgcaatc gccagtggca ctgctgtatc taaggtcctg 540
cacttagaag gagaagtgaa caagatcaaa agtgctctac tatccacaaa caaggccgta 600
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actgtgatag agttccaaca aaagaacaac agactactag agattaccag ggaatttagt 780
gttaatgcag gtgtaactac acctgtaagc acttacatgt taactaatag tgaattattg 840
tcattaatca atgatatgcc tataacaaat gatcagaaaa agttaatgtc caacaatgtt 900
caaatagtta gacagcaaag ttactctatc atgtccataa taaaagagga agtcttagca 960
tatgtagtac aattaccact atatggtgtg atagatacac cttgttggaa attacacaca 1020
tcccctctat gtacaaccaa cacaaaagaa gggtcaaaca tctgtttaac aagaactgac 1080
agaggatggt actgtgacaa tgcaggatca gtatctttct tcccacaagc tgaaacatgt 1140
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aaaacagatg taagcagctc cgttatcaca tctctaggag ccattgtgtc atgctatggc 1320
aaaactaaat gtacagcatc caataaaaat cgtggaatca taaagacatt ttctaacggg 1380
tgtgattatg catcaaataa aggggtagac actgtgtctg taggtaacac attatattat 1440
gtaaataagc aagaaggcaa aagtctctat gtaaaaggtg aaccaataat aaatttctat 1500
gacccattag tattcccctc tgatgaattt gatgcatcaa tatctcaagt caatgagaag 1560
attaaccaga gtttagcatt tattcgtaaa tccgatgaat tattacatca tgtaaatgct 1620
ggtaaatcaa ccacaaatta ataataatta agaaaaaata cgggtagaat cggagtgccc 1680
c 1681
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tccttaatat agctgaattg attgcagctg cgcgatc 37
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<211> Length : 37
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Sequence
5
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<400> PreSequenceString :
cagctatatt aagga 15
<212> Type : DNA
<211> Length : 15
SequenceName : 6
SequenceDescription :
Sequence
--------
<213> OrganismName :
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tcggagtgcc ccgat 15
<212> Type : DNA
<211> Length : 15
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SequenceDescription :
Claims (4)
1. the heat-resisting vaccine strain of recombinant Newcastle disease of the respiratory syncystial virus F protein that expression signal peptide is replaced, it is characterised in that institute
The heat-resisting vaccine strain Classification And Nomenclature stated is rTS-tF/RSV, is preserved in Wuhan University's Chinese Typical Representative on April 19th, 2016 and cultivates
Thing preservation center, preserving number is CCTCC NO:V201625.
2. the preparation method of the heat-resisting vaccine strain of recombinant Newcastle disease of the respiratory syncystial virus F protein that expression signal peptide is replaced, its
It is characterised by comprising the steps:
The amplification of the porcine circovirus 2 type Cap gene that 1.1 signal peptides are replaced
Being bred on Hep-2 cell by Respiratory Syncytial Virus Strain Long, after inoculating 5 days, multigelation harvesting is trained
Nutrient solution.The cell culture fluid of results is carried out the PCR amplification of truncate F gene, it is thus achieved that the F protein of truncate；With primer from extension
Method amplification obtains tPAs sequence；Truncate F gene order and tPAs sequence are attached by the way of over-lap PCR, then enter
Performing PCR expands, and amplimer is: forward primer 5 '-CAGCTATATTAAGGATTAAGAAAAAATACGGGTAGAAAGCTTGCC
ACCATGGATGC-3 ', downstream primer 5 '-GGGGCACTCCGATTCTACCCGTATTTTTTCTTAATTATTATTAATTTGTGG
TTG-3′；PCR primer carrying out agarose gel reclaim after purification, measure DNA concentration, pending clone connects；
The PCR amplification of 1.2 Avian pneumo-encephalitis virus heat-resistant carriers fragments
Obtaining Avian pneumo-encephalitis virus carrier sequence with the method for super LA-PCR, amplimer is: forward primer 5 '-GAATCGGAG
TGCCCCGATTGTGCCAAGATGGACTCATC-3 ', downstream primer 5 '-TCCTTAATATAGCTGAATTGATTGCAGCTGCGC
GATC-3′；Amplification template is Avian pneumo-encephalitis virus heat-resisting strain plasmid pTS09-C；Carry out PCR primer reclaiming purification, measure it dense
Degree, treats that clone connects；
The clone of 1.3 recombiant plasmid pTS-tF/RSV connects
Use In-fusion to clone interconnection technique, truncate F genetic fragment good for purification and heat-resistant carriers fragment are carried out clone even
Connecing, catenation sequence is CAGCTATATTAAGGA and TCGGAGTGCCCCGAT；Connect product and convert DH5 α competent cell, convert
After cell cultivate 16 hours on LB plate after choose single bacterium colony and carry out liquid culture, be accredited as the bacterium solution of the positive and carry out plasmid and carry
Take；
1.4 recombinant virus rTS-tF/RSV rescues recover
Use lipofection, the recombiant plasmid pTS-tF/RSV built and three are expressed NP, P and L albumen respectively
Helper plasmid cotransfection BHK-21 cell；Transfect latter 72 hours, multigelation harvesting liquid, inoculate 9-11 age in days SPF Embryo Gallus domesticus.
Inoculating latter 5 days, gather in the crops chick embryo allantoic liquid, the restructuring of the respiratory syncystial virus F protein that isolated expression signal peptide is replaced is new
The heat-resisting vaccine strain of city epidemic disease.
3. the heat-resisting vaccine of recombinant Newcastle disease of the respiratory syncystial virus F protein that the expression signal peptide described in claim 1 is replaced
Strain application in preparing recombinant virus.
4. the heat-resisting vaccine of recombinant Newcastle disease of the respiratory syncystial virus F protein that the expression signal peptide described in claim 1 is replaced
Strain application in preparing the heat-resisting live vaccine of respiratory syncytial virus sub-units.
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Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN113462660A (en) * | 2021-07-22 | 2021-10-01 | 武汉大学 | Recombinant Newcastle disease vector vaccine for expressing avian infectious bronchitis virus S protein, preparation method and application |
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| CN113462660A (en) * | 2021-07-22 | 2021-10-01 | 武汉大学 | Recombinant Newcastle disease vector vaccine for expressing avian infectious bronchitis virus S protein, preparation method and application |
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