CN1408434A - Polypeptide and DNA combined vaccine for resisting animal's foot-and-mouth disease and its preparing method - Google Patents
Polypeptide and DNA combined vaccine for resisting animal's foot-and-mouth disease and its preparing method Download PDFInfo
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Abstract
The combined vaccine consists of a kind of polypeptide vaccine and one DNA vaccine, and may be used to immunize suscaptible animal in several modes to prevent foot-mouth disease virus infection. The chemically synthetic encoding foot-mouth disease virus VP1 protein has the DNA sequence of antigen amino acid peptide segment and is connected into multicopy serial structure. The antigen gene with serial structure is then connected to beta-galactosidase gene to form fusion gene. When the fusion gene is inserted into protokaryon expression vector, the expressed fusion protein may be compounded into the polypeptide vaccine. When the fusion gene is inserted into eukaryon expression vector, the recombinant eukaryon expression plasmid may be amplified to obtain NDA vaccine.
Description
Technical field
The invention belongs to field of genetic engineering, be specifically related to a kind of polypeptide of anti-livestock foot-and-mouth disease and DNA combined vaccine and preparation method thereof.
Background technology
Foot and mouth disease (foot-and-mouth disease, FMD) be one of livestock contagious disease the most serious in the world today, cloven-hoofed animals such as main harm pig, cattle, sheep, and the outburst scope is wide, propagate rapidly, the mortality rate height causes enormous economic loss to animal husbandry behind the zoogenetic infection, thereby is classified as first of the category-A infectious disease by World Organization for Animal Health.
Advantages such as gene engineering polypeptide vaccine is easy to preserve because safety is good, and is satisfactory for result have broad application prospects.Yet necessary secondary immunity is the immune system of excitating organism better, makes it have enough resistances to foot and mouth disease virus (FMDV), thereby has increased the input of manpower and materials.For this reason, being necessary to seek more simple and effective method overcomes the above problems.
Dna vaccination is described as the revolution for the third time of vaccine biotechnology, and relevant work progress is rapid.The preparation method of dna vaccination is normally inserted carrier for expression of eukaryon with antigen gene, constitutes the eukaryon expression plasmid of reorganization.This plasmid has eucaryon replicon and promoter, can be in eukaryotic cell the relevant antigen of continuous expression, and then stimulate body to produce to continue antigen-specific immune responses.The shortcoming of dna vaccination is that the antigen presentation amount is not high, and immunne response is comparatively slowly with faint.
Summary of the invention
The objective of the invention is anti-livestock foot-and-mouth disease recombinant vaccine that proposes a kind of high-efficient and lasting and preparation method thereof,, bring into play the advantage of above-mentioned vaccine to overcome the deficiency of above-mentioned vaccine.
The anti-livestock foot-and-mouth disease recombinant vaccine that the present invention proposes, the combined vaccine of forming by polypeptide vaccine and dna vaccination, wherein, the DNA sequence that has antigenic amino acid peptide section in the chemical synthesis coding foot and mouth disease virus VP1 albumen, form the antigen polypeptide gene by fixed structure series connection, and be connected with beta-galactosidase gene, constitute fusion gene; If this fusion gene cloning to prokaryotic expression carrier, is expressed the fusion rotein that obtains certain molecular weight, will can obtain the aforementioned polypeptides vaccine after the fusion rotein emulsifying; If this fusion gene cloning to carrier for expression of eukaryon, is above-mentioned dna vaccination after the recombiant plasmid amplification.
Among the present invention, polypeptide vaccine is to be carrier protein with the beta galactosidase, and the foot-and-mouth disease virus antigen polypeptide chain is connected on the appropriate location of beta galactosidase, constitutes a fusion rotein.Wherein the FMDV antigen polypeptide adopts in the foot and mouth disease virus VP1 albumen and has antigenic amino acid peptide section, and with its cascaded structure of forming multicopy, adds suitable amino acid polypeptide as joint in peptide section junction.
Can be discerned and submission by the animal body immune system better in order to guarantee fusion rotein, carrier protein as this fusion rotein can be the suitable section of part or the full length sequence of beta galactosidase, and can be beta galactosidase from the 1st to the 583rd the homologous sequence of totally 583 amino acid peptide sections.1024 aminoacid of beta galactosidase total length, different colibacillary beta galactosidase nucleotide homologies are greater than 90%, and amino acid identity is greater than 95%.For the antigen polypeptide that guarantees to insert can embody antigenicity better, can choose different antigen and insert the site, the foot-and-mouth disease virus antigen polypeptide that will have cascaded structure inserts wherein.And antigen polypeptide can be connected the appropriate location of carrier protein beta galactosidase, inserts as N end, C end or in the centre position.
Among the present invention, antigen polypeptide adopts in the foot and mouth disease virus VP1 albumen has antigenic amino acid peptide section, adopts the multicopy cascaded structure simultaneously.For having antigenic amino acid peptide section, as O type foot and mouth disease virus VP1 protein 21 ヘ 40,141 ヘ 160,200 ヘ 213 amino acids peptide sections, and can before and after the float amino acid residue of suitable number, as adopting amino acid peptide section such as 209 of 35 ヘ, 53,134 ヘ, 158,141 ヘ, 158,135 ヘ, 144,188 ヘ; Aminoacid sequence in the FMDV epitope partly has conservative, and as the RGD site among the 141 ヘ 160AA, but not conservative part is a foundation with the epidemic isolates gene order; Usually as among the 21 ヘ 40AA have 1-4 aminoacid to be replaced, have 1-5 aminoacid to be replaced among the 141 ヘ 160AA, have 1-2 aminoacid to be replaced among the 200 ヘ 213AA, its principle is that these amino acid peptides Duan Jun has antigenicity.And,, also can adopt the cascaded structure of other multicopies (as two copies, four copies etc.) as the three copy cascaded structures of 141 ヘ 160AA-21 ヘ 40AA (or 200 ヘ 213AA)-141 ヘ 160AA for the cascaded structure of antigen polypeptide.In the cascaded structure, having the employed copy number of antigenic a certain amino acid peptide section can be different, and residing position can be different; When having antigenic amino acid peptide section and being connected to form cascaded structure, add suitable amino acid polypeptide as joint in peptide section junction, the aminoacid kind number in the joint can change as required.Its principle is can show enough antigenicities after forming fusion rotein with beta galactosidase.
Among the embodiment foot-and-mouth disease virus antigen polypeptide chain is connected on the C end of beta galactosidase, constitutes a fusion rotein.After making antigen polypeptide be expressed as fusion rotein, can embody antigenicity better, we have suitably chosen a certain section in the middle of the beta galactosidase as carrier protein, adopt among the embodiment as described later beta galactosidase since about 600 the amino acid residue peptide sections of the 1st amino acids residue as carrier protein; The FMDV antigen polypeptide then adopts 3 major antigen epi-positions in the O type foot and mouth disease virus VP1 albumen, i.e. 213 3 amino acid peptide sections of 21 ヘ 40,141 ヘ, 160,200 ヘ, when making up polypeptide vaccine, it is formed the cascaded structure of 141 ヘ 160AA-200 ヘ 213AA-141 ヘ 160AA; And when the structure dna vaccination, it is formed the cascaded structure of 141 ヘ 160AA-21 ヘ 40AA-141 ヘ 160AA, add suitable amino acid polypeptide as joint in peptide section junction.Adopt the antigen polypeptide structure of 141 ヘ 160AA (" Y ")-Pro-Gly (" M ")-21 ヘ 40AA (" X ")-Gln-Phe-Glu-Leu-Glu-Phe-Met-Val-Pro-Ser-Arg (" N ")-141 ヘ 160AA (" Y ") as described later among the embodiment, former and later two joints are respectively 2 and 11 aminoacid.
The polypeptide vaccine that the present invention relates to, adopt the FMDV antigen gene sequences with cascaded structure to be in an embodiment: SEQ ID NO.1, the antigen polypeptide aminoacid sequence of its coding are SEQ ID NO.2.
Its architectural feature is as follows:
Respectively there is the DNA sequence of coding 141 ヘ 160 amino acids of a copy at the two ends of antigen gene, and the centre is the DNA sequence of coding 200 ヘ 213 amino acids of a copy; Two joints that connect these sequences are respectively 2 and 11 aminoacid.
On gene level, this cascaded structure is connected the C-terminal of beta galactosidase, forms fusion gene, express obtaining fusion rotein.This fusion rotein has following characteristics: (1) on gene level, carrier protein gene size is about 1.8kb, and antigenic peptides gene size is about 200bp, and whole fusion gene size is about 2kb.(2) on protein level, carrier protein only comprises beta galactosidase since about 600 amino acids peptide sections of the 1st; Antigen protein is above-mentioned tandem polypeptide section, and wherein 141 ヘ 160 and 200 ヘ, 213 amino acids peptide Duan Jun are epitopes of B cell.Whole fusion protein molecule amount is about 72kDa.
The dna vaccination that the present invention relates to is to make up to form on the basis of carrier for expression of eukaryon.This eukaryon expression plasmid has the protokaryon replicon simultaneously, guarantees amplification in a large number in escherichia coli, as pcDNA, pCMV and other serial carrier for expression of eukaryon.The pCDM8-FZ2 eukaryon expression plasmid that makes up among the embodiment has the M13 replicon when having SV40 replicon and CMV promoter.Have pig immune globulin IgG signal peptide gene before the cloning site of this plasmid, guarantee that the antigen of expressing can be transported to outside the born of the same parents fully to contact in eukaryotic cell with immune system.In this plasmid, be connected with the described fusion gene of claim 1 at the signal peptide gene end, this fusion gene is made up of vector gene (being beta galactosidase) and antigen gene, and used in vector gene and the polypeptide vaccine to be used for the proteic beta-galactosidase gene of expression vector in full accord; Antigen gene then can be different from the antigen gene in the polypeptide vaccine.In the fusion gene, the vector gene that we adopt is consistent with the fusion gene in the polypeptide vaccine with the antigen gene connected mode.In order to make dna vaccination can excite the animal body immune system more effectively, we have added the CpG immunostimulatory sequence in eukaryon expression plasmid.
In an embodiment, the cascaded structure that antigen gene had that is used to make up eukaryon expression plasmid is 141 ヘ 160AA-21 ヘ 40AA-141 ヘ 160AA, and middle joint and the polypeptide vaccine that adopts is in full accord; This antigen gene sequences is: SEQ ID NO.3, the antigen polypeptide aminoacid sequence of its coding are SEQ ID NO.4.
The invention allows for the preparation method of above-mentioned combined vaccine.Comprise technology and methods such as adopting gene preparation, reorganization, the extensive extracting of plasmid and Expression of Fusion Protein, concrete steps are as follows:
(1) with the DNA sequence and the linker DNA sequence that have antigenic amino acid peptide section in the chemical synthesis process composite coding foot and mouth disease virus VP1 albumen, on dna level, connect into complete antigen gene, this gene is linked to each other with beta-galactosidase gene, constitute fusion gene;
(2) above-mentioned fusion gene is inserted prokaryotic expression carrier, and transformed into escherichia coli, express obtaining fusion rotein.This fusion rotein obtains the aforementioned polypeptides vaccine through preparation work such as Over emulsfications;
(3) above-mentioned fusion gene is inserted carrier for expression of eukaryon, and transformed into escherichia coli, the eukaryon expression plasmid of reorganization is bred in escherichia coli in a large number.Extensive this plasmid of extracting obtains above-mentioned dna vaccination through corresponding preparation work again.
The invention still further relates to the using method of this combined vaccine.Traditional vaccine and existing most of recombinant vaccine in use must carry out secondary immunity, just can make animal body have the infection of enough immunity opposing virus.Its advantage of combined vaccine of the present invention is to carry out once immunity can make animal body produce enough immunity, and lasting medicine, has fully overcome the deficiency of other vaccine many aspects.
The new generation vaccine that the present invention proposes is used to prevent dissimilar foot and mouth disease according to what make up according to difference.Consider the immunological effect of vaccine, we propose: with O type FMDV gene order serves as to be best suited for prevention O type foot and mouth disease according to the vaccine that makes up; With A type FMDV gene order serves as to be best suited for prevention A type foot and mouth disease according to the vaccine that makes up; In like manner, serve as to be best suited for according to the vaccine that makes up to prevent the foot and mouth disease of type separately with various FMDV gene orders such as AsiaI.Simultaneously, we also propose: the FMDV gene order with a certain strain of a certain type is the different strains that are widely used in susceptible animal prevention homotype foot and mouth disease virus according to the vaccine that makes up, reason is that variation is less between the different strains of homotype foot and mouth disease virus, with the animal of this vaccine immunity these strains is had cross immunity power.
The novel combined vaccine that the present invention is proposed carries out the immune efficacy check.Attack with O type foot and mouth disease virus behind the certain hour not by the pig of natural infection so that a certain amount of this combined vaccine is once immune, the result shows that this vaccine has very good protection effect to laboratory animal, and protective rate reaches 100% (seeing Table 1).
The immune protective effect vaccine immunizing dose virus attack dosage of table 1 pair pig is attacked pig's head number morbidity pig's head and is counted protective rate immune group 4mg peptide Seedling associating 100ID
50/ 2ml 50 100%
1mgDNA vaccine matched group 0 100ID
50/ 2ml 550
The pluses and minuses of comprehensive gene engineering polypeptide vaccine of the present invention and dna vaccination have proposed a kind of foot-and-mouth disease virus resistant combined vaccine and preparation and using method, promptly adopt the infection of immune animal of polypeptide vaccine and dna vaccination associating with the prevention foot and mouth disease virus.In this combined vaccine, on the one hand, the existence of polypeptide vaccine can be brought out body and produce antigen-specific immune responses apace; On the other hand, the existence of dna vaccination can make immune system obtain relevant antigenic long stimulus, and foot and mouth disease virus is kept long resistance.
The specific embodiment
Be that example further specifically describes the present invention with Schweineseuche O-shaped polypeptide and DNA combined vaccine below.
The building process of polypeptide vaccine.With coding 200 ヘ segmental DNA sequence of 213,141 ヘ, 160 amino acids and linker DNA sequences in the synthetic VP1 gene of chemical method, series connection becomes the primary structure (SEQ ID NO.1) of 141 ヘ 160AA-200 ヘ 213AA-141 ヘ 160AA on gene level, and is inserted into cloning vehicle pBluescriptIIKS.Get 1ug pBluescriptIIKS with EcoR I and BamH I double digestion, in 37 ℃ of reactions two hours, big fragment was reclaimed in the electrophoresis rubber tapping.This large fragment DNA is mixed with the above-mentioned fragment of connecting, under effects such as T4 ligase, coupled reaction takes place.The recombinant DNA transformed into escherichia coli competent cell that obtains is incubated in the ampicillin flat board, and 37 ℃ of inversions are spent the night.Picking white transformant with BstEII single endonuclease digestion, EcoR I and BamH I double digestion, pcr amplification, dna sequencing Analysis and Identification transformant, thereby obtains positive colony.With EcoR I and two above-mentioned positive colony, the electrophoresis rubber tapping recovery small fragments cut of BamH I.With the two expression plasmid carrier pWR590 that cut of these two restricted enzyme, big fragment is reclaimed in the electrophoresis rubber tapping equally.In like manner be connected, transform and screen with above-mentioned, finally obtain positive colony, called after pXZ500.
The building process of dna vaccination.With coding 21 ヘ segmental DNA sequence of 40,141 ヘ, 160 amino acids and linker DNA sequences in the synthetic VP1 gene of chemical method, series connection becomes the primary structure (SEQ ID NO.3) of 141 ヘ 160AA-21 ヘ 40AA-141 ヘ 160AA on gene level, and with this antigen gene formation fusion gene that links to each other with beta galactosidase.Fusion gene cloning is arrived carrier for expression of eukaryon pCDM8.Method similar to the above is identified transformed clone.The positive colony called after pCDM8-FZ2 that obtains.
Is fermentation liquid with pXZ500 and pCDM8-FZ2 with the LB culture fluid, 37 ℃ of mixing speed 6000rpm aerobic culture.PXZ500 collects thalline after cultivating 12 hours, thalline is suspended in the shell-broken liquid Ultrasound Instrument breaking cellular wall of usefulness 95W power 5 minutes again; Ultrasonic back 5000rpm collected antigen protein in centrifugal 20 minutes, it is made into oil emulsion can obtains polypeptide vaccine in claims.The pCDM8-FZ2 overnight incubation is also collected thalline, with the extensive extracting plasmid DNA of alkaline process, can obtain the dna vaccination in claims after the dissolving.
At last above-mentioned two kinds of vaccines are hybridly prepared into anti-foot and mouth disease polypeptide and DNA combined vaccine, are used for immune foot and mouth disease susceptible animal.
Relevant animal disease resistant poison ability test experience the results are shown in Table 1.
The sequence that the present invention relates to is as follows:
SEQ ID NO.1：
GTA?CCA?AAC?CTG?CGT?GGT?GAC?CTG?CAG?GTA?CTT?GCT?CAG?AAA?GTT?GCT?CGT?ACT
CAT?GGT?TTG?GAC?GCA?CCA?CTG?GAC?GTC?CAT?GAA?CGA?GTC?TTT?CAA?CGA?GCA?TGA
CTG?CCA?CCC?GGG?CGT?CAC?AAA?CAG?AAA?ATC?GTA?GCT?CCA?GTA?AAA?CAG?ACT?CTGGAC?GGT?GGG?CCC?GCA?GTG?TTT?GTC?TTT?TAG?CAT?CGA?GGT?CAT?TTT?GTC?TGA?GACCAA?TTC?GAG?CTC?GAA?TTC?ATG?GTA?CCC?TCG?AGG?GTA?CCA?AAC?CTG?CGT?GGT?GACGTT?AAG?CTC?GAG?CTT?AAG?TAC?CAT?GGG?AGC?TCC?CAT?GGT?TTG?GAC?GCA?CCA?CTGCTG?CAG?GTA?CTT?GCT?CAG?AAA?GTT?GCT?CGT?ACT?CTG?CCAGAC?GTC?CAT?GAA?CGA?GTC?TTT?CAA?CGA?GCA?TGA?GAC?GGTSEQ?ID?NO.2：VPNLRGDLQVLAQKVARTLPPGRHKQKIVAPVKQTLQFELEFMVPSRVPNLRGDLQVLAQKVARTLPSEQ?ID?NO.3：GTG?AGC?AAC?GTG?AGG?GGT?GAC?CTT?CAA?GTG?TTG?GCT?CAG?AAG?GCA?GAA?AGA?GTTCAC?TCG?TTG?CAC?TCC?CCA?CTG?GAA?GTT?CAC?AAC?CGA?GTC?TTC?CGT?CTT?TCT?CAACTG?CCC?CCC?GGT?GAG?ACA?CAG?GTC?CAG?AGA?CGC?CAG?CAC?ACG?GAT?ATC?TCG?TTTGAC?GGG?GGG?CCA?CTC?TGT?GTC?CAG?GTC?TCT?GCG?GTC?GTG?TGC?CTA?TAG?AGC?AAAATA?CTA?GAC?AGA?TTT?GTG?CAG?TTT?GAG?CTG?GAG?TTT?ATG?GTG?CCC?AGC?AGG?GTGTAT?GAT?CTG?TCT?AAA?CAC?GTC?AAA?CTC?GAC?CTC?AAA?TAC?CAC?GGG?TCG?TCC?CACAGC?AAC?GTG?AGG?GGT?GAC?CTT?CAA?GTG?TTG?GCT?CAG?AAG?GCA?GAA?AGA?GTT?CTGTCG?TTG?CAC?TCC?CCA?CTG?GAA?GTT?CAC?AAC?CGA?GTC?TTC?CGT?CTT?TCT?CAA?GACCCCGGGSEQ?ID?NO.4：VSNVRGDLQVLAQKAERVLPPGETQVQRRQHTDISFILDRFVQFELEFMVPSRVSNVRGDLQVLAQKAERVLP
Claims (13)
1, a kind of foot and mouth disease gene vaccine, it is characterized in that uniting group by polypeptide vaccine and dna vaccination, wherein, the DNA sequence that has antigenic amino acid peptide section in the chemical synthesis coding foot and mouth disease virus VP1 albumen, form the antigen polypeptide gene by fixed structure series connection, and be connected with beta-galactosidase gene, constitute fusion gene; If this fusion gene cloning to prokaryotic expression carrier, is expressed the fusion rotein that obtains certain molecular weight, will can obtain the aforementioned polypeptides vaccine after the fusion rotein emulsifying; If this fusion gene cloning to carrier for expression of eukaryon, is above-mentioned dna vaccination after the recombiant plasmid amplification.
2, recombinant vaccine according to claim 1, the carrier protein that it is characterized in that fusion rotein be beta galactosidase from the 1st to the 583rd totally 583 amino acid peptide sections.
3, recombinant vaccine according to claim 1 and 2, the carrier protein that it is characterized in that fusion rotein be beta galactosidase from the 1st to the 583rd the homologous sequence of totally 583 amino acid peptide sections.
4, recombinant vaccine according to claim 1, it is characterized in that antigen polypeptide adopts following cascaded structure: Y-M-X-N-Y or Y-M-Z-N-Y, wherein, " X " of same cascaded structure, " Y ", " Z " peptide section derives from same type foot and mouth disease virus VP1 protein sequence, and " X ", " Y ", " Z " peptide section is represented foot and mouth disease O type virus VP 1 protein 21 ヘ 40 respectively, 141 ヘ 160,200 ヘ, 213 amino acids peptide sections, perhaps foot and mouth disease A, the corresponding amino acid peptide section of AsiaI type virus surface proteins, " M " is " Pro-Gly " two peptide linkers, and " N " is " Gln-Phe-Glu-Leu-Glu-Phe-Met-Val-Pro-Ser-Arg " 11 peptide linkers.
5, recombinant vaccine according to claim 4, it is characterized in that " X " in the antigen polypeptide, " Y ", " Z " peptide section can before and after the float amino acid residue of suitable number: adopt 35 ヘ, 53,134 ヘ, 158,141 ヘ, 158,135 ヘ, 144,188 ヘ, 209 amino acids peptide sections.
6, according to claim 4 or 5 described recombinant vaccines, the aminoacid that it is characterized in that doing in " X " in the former polypeptide, " Y ", " Z " peptide section can be adjusted because of the needs that prevent different foot and mouth disease variation strains: have 1-4 aminoacid to be replaced in " X " peptide section, there is 1-5 aminoacid to be replaced in " Y " peptide section, has 1-2 aminoacid to be replaced in " Z " peptide section.
7, gene engineering vaccine according to claim 1 is characterized in that being used to prepare the used eukaryon expression plasmid of dna vaccination has following feature:
(1) makes up the used carrier for expression of eukaryon of this eukaryon expression plasmid and have the protokaryon replicon simultaneously;
(3) this plasmid has immunostimulatory sequence;
(2) this plasmid has signal peptide gene and eukaryotic promoter;
(4) in this plasmid, be connected with the fusion gene described in the claim 1 at the signal peptide gene end.
8, recombinant vaccine according to claim 7, the antigen gene that it is characterized in that being used in the dna vaccination making up the eukaryon expression plasmid fusion gene can be different from the antigen gene in the polypeptide vaccine, but all must derive from same type foot and mouth disease virus VP1 protein sequence.
9, recombinant vaccine according to claim 1 is characterized in that the antigen polypeptide of polypeptide vaccine is connected the C end of carrier protein.
10, according to the described recombinant vaccine of one of claim 1-9, it is characterized in that the antigen polypeptide gene order is: SEQ ID NO.1, the antigen polypeptide aminoacid sequence of its coding is SEQ ID NO.2, and SEQ ID NO.3, the antigen polypeptide aminoacid sequence of its coding are SEQ ID NO.4.
11, the application of recombinant vaccine as claimed in claim 1 is characterized in that with a certain type foot-and-mouth disease virus gene sequence serving as to be applicable to this type foot and mouth disease of prevention according to the vaccine that makes up; And have cross-protection: the FMDV gene order with a certain strain of a certain type is the infection that extensively is best suited for the different strains of susceptible animal prevention homotype foot and mouth disease virus according to the vaccine that makes up.
12, the preparation method of recombinant vaccine as claimed in claim 1 comprises gene preparation, reorganization, the extensive extracting of plasmid and Expression of Fusion Protein etc., it is characterized in that concrete steps are as follows:
(1) with the DNA sequence that has antigenic amino acid polypeptide in the chemical synthesis process composite coding foot and mouth disease virus VP1 albumen, on dna level, connects into complete antigen gene, this gene is linked to each other with beta-galactosidase gene, constitute fusion gene;
(2) above-mentioned fusion gene is inserted prokaryotic expression carrier, and transformed into escherichia coli, express obtaining fusion rotein.This fusion rotein obtains the aforementioned polypeptides vaccine through preparation work such as Over emulsfications;
(3) above-mentioned fusion gene is inserted carrier for expression of eukaryon, and transformed into escherichia coli, extensive this plasmid of extracting obtains above-mentioned dna vaccination through corresponding preparation work again.
13, the using method of recombinant vaccine as claimed in claim 1 is characterized in that polypeptide vaccine and dna vaccination mix or immunity respectively; But according to the immunity of dosage one or many.
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Cited By (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN100422326C (en) * | 2004-08-26 | 2008-10-01 | 复旦大学 | Si RNA capable of controlling foot-and-mouth disease virus copying and infection and its preparing method and use |
| CN103255159A (en) * | 2013-03-04 | 2013-08-21 | 张掖市奥林贝尔生物科技有限公司 | Recombinant expression and preparation method of cecropin CAD |
| CN105209620A (en) * | 2013-03-15 | 2015-12-30 | 诺瓦瓦克斯股份有限公司 | Enhanced expression of picornavirus proteins |
| CN109196104A (en) * | 2016-04-11 | 2019-01-11 | 生源霸科乌拉圭有限公司 | Wide acting type vaccine for viral disease |
| CN110382518A (en) * | 2016-10-31 | 2019-10-25 | 美国农业部 | Chimeric for serotypes A type foot and mouth disease virus |
-
2002
- 2002-09-16 CN CN 02137010 patent/CN1408434A/en active Pending
Cited By (8)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN100422326C (en) * | 2004-08-26 | 2008-10-01 | 复旦大学 | Si RNA capable of controlling foot-and-mouth disease virus copying and infection and its preparing method and use |
| CN103255159A (en) * | 2013-03-04 | 2013-08-21 | 张掖市奥林贝尔生物科技有限公司 | Recombinant expression and preparation method of cecropin CAD |
| CN103255159B (en) * | 2013-03-04 | 2015-11-18 | 张掖市奥林贝尔生物科技有限公司 | A kind of cecropin CAD's is recombinant expressed and preparation method thereof |
| CN105209620A (en) * | 2013-03-15 | 2015-12-30 | 诺瓦瓦克斯股份有限公司 | Enhanced expression of picornavirus proteins |
| CN109196104A (en) * | 2016-04-11 | 2019-01-11 | 生源霸科乌拉圭有限公司 | Wide acting type vaccine for viral disease |
| CN109196104B (en) * | 2016-04-11 | 2023-01-31 | 生源霸科乌拉圭有限公司 | Broad-spectrum vaccine for viral diseases |
| CN110382518A (en) * | 2016-10-31 | 2019-10-25 | 美国农业部 | Chimeric for serotypes A type foot and mouth disease virus |
| CN110382518B (en) * | 2016-10-31 | 2022-11-29 | 美国农业部 | Chimeric vaccine for serotype A foot and mouth disease virus |
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