78 results about "Vp2 gene" patented technology

The invention relates to construction and application of recombinant nucleocapsids of Cap genes of porcine circovirus expression type 2 nucleocapsid protein, belonging to the genetic engineering bacterin field. C-terminal gene fragments of porcine parvovirus (PPV) VP2 genes are cloned into a type 5 adenovirus shuttle vector of the human beings, and recombinant adenovirus rAd-deltaVP2 is obtained; deltaVP2 proteins are expressed successfully and highly efficiently and can be self-assembled into the nucleocapsids [PPV:VLPs]; the PPV VP2 nucleocapsids are used as antigen transport vectors and 165 to 200 sites of amino acid (deltaCap) genes of the porcine circovirus type 2 (PCV2) nucleocapsid proteins (Cap) are embedded into an N-terminal (deltaVP2) of the PPV VP2, and then recombinant adenovirus rAd-deltaCap-deltaVP2 is obtained; embedded VP2 (deltaCap-deltaVP2) proteins are expressed successfully and highly efficiently and can be self-assembled into nucleocapsids [PPV:VLP(PCV2)]. The invention also relates to application of the recombinant virus and recombinant PPV VP2 nucleocapsids of the expression Cap genes of the recombinant virus in the aspects of bacterin immunity and so on.

The invention discloses immune colloidal gold test paper capable of simultaneously detecting canine distemper and canine parvo of foxes, raccoon dogs and minks and a method for preparing the immune colloidal gold test paper. The test paper consists of a test paper box and a test paper tape, and the test paper tape is made by sequentially bonding a hard polyvinyl chloride lining plate, a nitrocellulose membrane, a long colloidal gold combination pad, a short colloidal gold combination pad, a sample pad and an absorption pad. VP2 gene monoclonal antibody- colloidal gold biomarkers resistant to mink-source canine distemper viruses and parvoviruses are sprayed on the long and short colloidal gold combination pads respectively. Two detection lines formed by rabbit anti-mink source canine distemper virus and parvovirus antibodies and a quality control line formed by a rabbit antimouse antibody are sprayed on the nitrocellulose membrane. The test paper is applicable to quick detection of pestilences of foxes, raccoon dogs and minks, and detection of two pestilences including canine distemper and canine parvo of foxes, raccoon dogs and minks can be simultaneously completed by the same test paper, and detection is simple, convenient, quick and accurate.

The invention relates to virus-like particle recombinant protein of a virus variation strain VP2 gene of an infectious bursal disease, belonging to the field of biologic pharmacy. The IBDV variation strain (AH1) VP2 gene is cloned, converted and transfected to obtain a recombinant baculovirus vBac-VP2; an infected Sf9 insect cell has specific fluorescence, and the antigen valence of the infected Sf9 insect cell is above 1.6*10<3>; the molecular weight of a recombinant VP2 protein is 53kDa, and the recombinant VP2 protein is in the state of virus-like particles; an indirect ELISA detection method established for an envelope antigen by the purified recombinant VP2 protein has good specificity and sensibility; an immune chicken can resist IBDV virulent attack, and the protection ratio achieves 100 percent. The novel virus-like particle recombinant VP2 protein prepared by the IBDV variation strain VP2 gene has high pertinence on the immune prevention of a current prevalent IBDV virulent strain and good practical value and popularization prospects.

The invention provides an anti-chicken infectious bursal disease (IBD) recombinant protein subunit vaccine. The vaccine is a fusion protein having high immunogenicity of Salmonella typhimurium flagellin and an infectious bursal disease virus (VP2). The above flagellin + VP2 fusion protein is obtained through the expression of a recombinant baculovirus containing a flagellin + VP2 gene by utilizing a Bac-to-Bac baculovirus expression system. The recombinant baculovirus obtained through the system has a short period, and the expressed flagellin + VP2 fusion protein has high immune protection force.

The invention discloses a VP2 gene restructuring Newcastle disease LaSota weak virus strain rLasota-VP2 of Infectious bursal disease virus, IBDV and appliance in the Newcastal disease virus,NDV and IBDV vaccine.

The invention discloses a preparation method of an anti-canine parvovirus protein VP2 specific IgY, which comprises the following steps: (1) designing a pair of primers according to CPV-VP2 gene sequence, carrying out PCR (polymerase chain reaction) amplification on the CPV-VP2 gene, connecting to a pMD18-T vector, transforming DH5alpha competent cell, carrying out blue-white screening, extracting the plasmid, carrying out digest enzyme digestion ion analysis, carrying out positive plasmid sequencing, and carrying out comparative analysis on the sequencing result; (2) expression and purification of VP2 protein in Escherichia coli: carrying out BamH I and Xho I double-enzyme digestion on the pMD18-T-VP2 and pET-32a,connecting to the target segment, constructing the pET-32a-VP2 expression vector, transforming Bal21(DE3)pLysS competent bacteria, carrying out enzyme digestion and PCR identification, optimizing the IPTG (isopropyl-beta-D-thiogalactopyranoside) induction concentration and time, carrying out mass induction expression, and purifying the recombinant protein by using a Ni<+> affinity column; and (3) preparation of anti-VP2-IgY antibody: immunizing a laying hen by using the purified VP2 protein, extracting the specific IgY antibody by using PEG 6000, and carrying out SDS-PAGE (sodium dodecyl sulfate polyacrylamide gel electrophoresis) analysis. The anti-VP2-IgY antibody extracted by the method can be well combined with the VP2 protein to carry out non-cross reaction on the degradation segment.

The invention discloses a recombinant rabies virus rHEP-Flury(dG-VP2) carrying two genes G and a gene VP2. According to the recombinant virus, a rabies virus HEP-Flury strain serves as a framework, the VP2 gene shown as SEQ ID NO.1 is inserted into HEP-Flury, an additional gene G of the rabies virus is inserted, and the recombinant plasmid pHEP-Flury(dG-VP2) carrying the two genes G and the gene VP2 is obtained; finally, rescuing screening is carried out, and the recombinant rabies virus rHEP-Flury(dG-VP2) is obtained. The recombinant rabies virus rHEP-Flury(dG-VP2) carries the two genes G and expresses VP2 protein, a high rabies virus resisting antibody and canine parvovirus VP2 resisting antibody level can be generated through immune induction, the cost of vaccines for dogs can be reduced, and very good application and popularization prospects are achieved.

The present invention relates to one kind of recombinant Newcastle disease Lasota low virulent vaccine strain expressing infectious bursal disease virus (IBDV) VP2 gene, and is especially recombinant Newcastle disease Lasota low virulent vaccine strain rLasota-VP2. The present invention also discloses the preparation process of the recombinant Newcastle disease Lasota low virulent vaccine strain and the application of the recombinant Newcastle disease Lasota low virulent vaccine strain in preparing vaccine for preventing IBDV caused diseases.

The invention relates to a method for preparing gosling plague virus-like granules with an escherichia coli system for soluble expression of gosling plague virus VP2 protein. The method for soluble expression of gosling plague virus VP2 protein comprises the following steps: performing codon optimization on a gosling plague virus VP2 gene, performing site-specific mutagenesis, namely, mutating a codon AGA into CGC and mutating GGA into GGT, cloning to a pET-Sumo vector, establishing a recombinant expression vector pET-Sumo-VP2, transforming the pET-Sumo-VP2 into a prokaryotic expression bacterium, and inducing with IPTG (isopropyl beta-D-1-Thiogalactopyranoside) at 37 DEG C so as to obtain soluble recombinant VP2 recombinant protein; and performing digestion on the recombinant protein with a ULP enzyme, and purifying with a Ni column, thereby obtaining purified VP2 protein. Electron microscope results show that the gosling plague virus-like granules can be prepared from VP2 protein after digestion, and moreover, the purified VP2 protein has good reactogenicity and can be applied to preparation of subunit vaccines of gosling plague virus genetic engineering.

The invention belongs to the field of virus detection, and relates to primers and a probe sequence for fluorescence RT-PCR detection for bluetongue virus serum-16. A real-time fluorescence RT-PCR detection method for BTV-16 comprises the following steps: adopting a Taqman technology, designing a group of specific primers which are only conservative in BTV-16 VP2 genes and a specific fluorescence labelling probe, and applying a fluorescence PCR instrument to detect a sample. Domestic and overseas BTV-16 viral nucleic acids can be specifically detected by using the fluorescence RT-PCR detection method established by the group of primers and the probe, and other serotypes of BTV nucleic acids cannot be detected. The method has the characteristics of being high in specificity, high in sensitivity, simple to operate, low in time consumption, high in efficiency and the like, and is capable of avoiding possible environmental pollution during a detection process.

The invention discloses a universal PCR detection kit for rapid and high-efficient detection of cat and dog parvovirus. The method comprises designing a pair of primers in accordance with a conservation region of a vp2 gene of the cat and dog parvovirus, performing a PCR reaction by using excrement of detected animals as a template, comparing with negative and positive controls included in the kit, and determining whether the detected animals are infected with the parvovirus. The detection kit is simple, easy to use, high in sensitivity, and good in stability, and can be used for monitoring health conditions of the cat and the dog.

The invention relates to an infectious bursal disease VP2 gene, wherein the amino acid sequence of the gene is SEQ ID NO: 1. The infectious bursal disease VP2 gene sieved by the invention is used as a subunit vaccine in genetic engineering. The subunit vaccine prepared by the infectious bursal disease VP2 gene sieved by the invention makes SPF chickens which are 21 days old immune. After 21 days, chickens are of strong immunity, so that the immune chickens can resist the attack of virulent virus of infectious bursal disease.

The invention discloses a recombinant yeast strain for expressing chicken IBDV (infectious bursal disease virus) VLPs (virus-like particles), a protein expressed by the recombinant yeast strain and an application. The recombinant yeast strain contains an optimized chicken IBDV VP2 gene sequence and can efficiently express a VP2 protein, and the expressed recombinant VP2 protein can be self-assembled into the VLPs. A vaccine is prepared from the chicken IBDV VLP protein expressed by the recombinant yeast strain to immunize chickens, and an experiment result indicates that the subunit peptide vaccine can effectively induce organisms to produce specific humoral immune response, so that the immunized chickens acquire 100% resistance to highly pathogenic vv IBDV fatal attack, residual viruses in the organisms can be cleared, and the purpose of sterilizing immunity is achieved. Therefore, one novel technical means is provided for prevention and control of chicken infectious bursal disease.

The invention provides a mink parvovirus virus-like particle which is characterized in that VP2 gene of mink parvovirus is optimized according to insect cell optimal codons, the 5' end of the optimized VP2 gene is directly connected with the nucleotide sequence of polyhedron of a coding part and then is cloned into a transfer vector, and the mink parvovirus virus-like particle can be produced through a baculovirus/insect cell expression system. The mink parvovirus virus-like particle can be produced through the expression system in a safe, efficient and large-scale manner, and the obtained virus-like particle is high in titer and good in immunogenicity; both muscle immunization and oral immunization can induce a mink body to produce high-level specific antibody which can resist attack of virulent virus and well protect a mink, and a foundation is laid for the preparation of mink viral enteritis vaccine.

The invention discloses a recombined rhabdovirus for expressing a porcine bocavirus VP2 protein and application thereof. A porcine bocavirus VP2 gene is amplified by PCR (polymerase chain reaction), and the nucleotide sequence of the porcine bocavirus VP2 gene is represented by SEQ ID NO.1. The amplified porcine bocavirus VP2 gene is inserted into BamHI and XhoI locuses of a rhabdovirus transmissible plasmid pFastBacTMHTB to construct a recombined transmissible plasmid pFastBac HTB-VP2; the recombined transmissible plasmid is converted into a DH10Bac competent cell comprising a rhabdovirus framework plasmid; the VP2 gene is integrated into the rhabdovirus framework plasmid Bacmid through homologous recombination; an sf9 cell is transfected by extracting the DNA of the recombined Bacmid to obtain the recombined rhabdovirus rBV-VP2 of which the preservation number is CCTCC V201401. A large quantity of VP2 protein and virus sample particles are expressed and produced in insect cells, and an effective PBoV VLPs subunit vaccine is developed.

A polymer protein (VP2/VP4/VP3) gene of infectious Fabricius bursa virus (IBDV) for intensifying the immunogenicity of vaccine and a recombinant carrier using pCI as eukaryon expression vector and containing said coding sequence are disclosed. The expression ability of said recombinant carrier increase by 10-40 times than normal eukaryon carrier. A DNA vaccine for infectious Fabricius bursa virus is also disclosed, which contains said eukaryon expression plasmid and immunological adjuvant and features high safety, stability and effect.

The present invention provides an avian recombinant herpesvirus modified by the presence of the cDNA encoding, the VP2 of the Delaware Variant E strain of IBDV, a subtype of IBDV serotype 1 strains. The present invention further provides an avian recombinant herpesvirus comprised of the VP2 gene of which the backbone virus is a Marek's disease vaccine strain, such as herpesvirus of turkeys. A poultry vaccine including the avian herpes recombinant virus described in the present invention can induce in chickens protective immunity against a variety of different subtypes of IBDV.

The invention belongs to the technical field of biology and in particular discloses a recombinant herpesvirus of turkey (HVT). The recombinant HVT is capable of simultaneously expressing NDV and IBDVprotective antigen genes, and is a bacterial artificial chromosome (BAC) based on the HVT. The construction method comprises the following steps: inserting a VP2 gene expression cassette of an infectious bursal disease virus LX strain under regulation of a cytomegalovirus promoter CMV into a UL45-46 non-essential region of the HVT genome by utilizing a gene recombination technology; and insertingan F gene expression cassette of a newcastle diseases virus PX02/3 strain under regulation of a chicken beta-actin promoter pec into another non-essential region US1-10 of the HVT genome. The recombinant herpesvirus of turkey is a recombinant triplet poultry vaccine candidate strain capable of controlling Marek's disease of chickens and effectively resisting Newcastle disease and infectious bursaldisease.

The invention provides a recombinant Lactobacillus casei for expressing the infectious pancreas necrosis virus (IPNV) VP2 protein and a preparation method thereof. According to the whole gene sequence of the IPNV VP2 protein and the gene fusion characteristic of expression vector plasmid, a pair of primers are designed to perform PCR to obtain target segment of 1095bp containing the main antigen site of the IPNV VP2 protein, the IPNV VP2 protein obtained through amplification is linked to the surface expressed carrier plasmid pPG1 and enters the cell of the host strain Lactobacillus casei 393 through electrotransformation, and the Lactobacillus casei containing positive recombinant plasmid pPG1-IPNV VP2 is named pPG1-IPNV VP2/L. casei 393. The recombinant pPG1-IPNV VP2/L. casei 393 is expressed under the induction of lactose. In the invention, the Lactobacillus casei expression carrier system of the IPNV VP2 protein is constructed and the target protein of around 40 KDa containing the main antigen site of the IPNV VP2 protein is expressed, and according to the Western blot experiment and indirect immunofluorescence experiment, the expressed foreign protein is capable of reacting with the IPVN immune serum, which shows that the recombinant VP2 protein and the IPNV natural antigen have the same antigenicity.

The invention provides a subunit vaccine of PPV (porcine parvovirus) disease and a preparation method of the subunit vaccine. PPV virus-like particles are provided firstly and produced by steps as follows: optimized VP2 gene and IL-15 (interleukin-15) gene are subjected to fusion expression, and the PPV virus-like particles are produced by a baculovirus/insect cell expression system. The PPV virus-like particles are mixed with a preservative and an adjuvant, and the PPV disease subunit vaccine is prepared. The PPV disease vaccine has the advantages of high yield, low production cost, high immunogenicity, good safety and the like, facilitates large-scale production, and has good immunization and prevention effects on PPV disease.