169 results about "Hog cholera virus" patented technology

The invention discloses a multiplex real-time fluorescent quantitation PCR method capable of detecting porcine circovirus, porcine parvovirus, porcine pseudorabies virus and hog cholera virus at the same time. The method of the invention detects that the four virus are all in good linear relations at the same time, all ten times serial dilution points of constructed normal plasmid are all in one straight line, CT value and copy number are in good linear relation, and regression analysis shows that the related coefficient of the CT value and the copy number is R<2> more than 0.99.The multiplex real-time fluorescent quantitation PCR method has the advantages of excellent specificity, sensibility and stability, which can rapidly, sensitively and differentially detect the four virus with serious harm to the economy and can be used for early diagnosis of virus infection.

The invention discloses a method for preparing live vaccines of hog cholera and a product thereof. The preparation method comprises the following steps of: (1) culturing porcine passage cell lines; (2) inoculating the porcine passage cell lines with live vaccine production seed viruses of the hog cholera to obtain attenuated vaccine strains of the hog cholera; (3) performing virus multiplication on the attenuated vaccine strains of the hog cholera; (4) measuring the virus titer of multiplication virus suspension by adopting an immunofluorescence method; and (5) adding a freeze-drying protective agent and antibiotics into the virus suspension which is detected to be qualified for vaccine matching and freeze-drying. The preparation method has the advantages of producing the live vaccines of the hog cholera by using the cell lines so as to achieve small quality differences among batches and the characteristics of simple and stable process, easy operation, high yield, low cost, the feasibility and extendibility of industrial production and the like, and measuring the virus titer of the multiplication virus suspension by adopting the immunofluorescence method so as to achieve sensitive, fast, specific and accurate detection, high repeatability and reliable results. The live vaccines of the hog cholera prepared by the method can completely protect pigs from the attacks of violent hog cholera viruses.

The invention discloses a fluorescence quantitative PCR detection reagent for African swine fever virus, and a preparation method and the application thereof. A set of specific primers and Taqman probes are designed and synthesized to be used for detecting ASFV P54 in relevant porcine products. A standard curve drawn in the invention provides a standard for the quantitative detection of ASFV P54. The invention establishes a fast and simple real-time fluorescence quantitative PCR detection system with strong specificity and high flexibility. The detection time is only several hours, and the detection lower limit can be 15 copies. The invention can be applied to the diagnosis and quarantine technology towards the imported relevant porcine products at port, and the invention provides reliable and effective technical condition for the import quarantine work of the country without ASF.

The invention discloses a reagent, detection method and application for the detection of African hog cholera virus. The reagent comprises a specific primer pair and a probe, upstream and downstream primers of the primer pair are respectively sequences shown in Seq ID No. 1 and 2, and the probe is a sequence shown in Seq ID No. 3 or its reverse complement sequence; in the probe sequence shown in the SEQ ID No.3, a 28th base modifies fluorescent quenching group-dT, a 31st base is replaced with a base analog, a 33rd base modifies a fluorescent group-dT, and a 3' end modifies C3 Spacer. The reagent is sensitive, specific and efficient for detection of the African hog cholera virus by recombinase polymerase amplification; compared with conventional conventional or real-time fluorescent PCR, thereagent and the detection method have short detection time and simple operation, are especially suitable for on-site testing, and are of great significance for the rapid prevention and control of theAfrican hog cholera virus and guarantee of production safety.

The invention belongs to the technical fields of immunoassay and biosensing and discloses a preparation method and application of an electrochemical immunosensor based on an HS-beta-CD-Ag-GOD conjugate. The electrochemical immunosensor is used for rapidly detecting a hog cholera virus antigen CSFV. The manufacturing scheme comprises the following steps: modifying a working electrode a bare platinum carbon electrode by using MWCNTs-CD-Fc-Ab1, and sequentially adding bovine serum albumin, hog cholera virus antigen CSFV and an Ab2-HS-beta-CD-Ag-GOD conjugate. The HS-beta-CD-Ag-GOD conjugate can convert glucose into gluconic acid, two protons and two electrons are transferred to oxygen molecules, hydrogen peroxide is generated, and an HS-beta-CD-Ag nanometer material serves as a mimic enzyme, the reduction of the hydrogen peroxide is catalyzed, the dual amplification of an electrochemical signal is realized, the high sensitivity is realized, and the detection limit can be low to 0.33pg / mL.

The invention discloses an RT-RPA (reverse transcription recombinase polymerase amplification) detection kit for fast detecting a high-pathogenicity porcine reproductive and respiratory syndrome virus and application thereof. The kit comprises a pair of primers and a probe, the sequences of the primers are shown as SEQ ID NO.1 and SEQ ID NO.2, and the sequence of the probe is shown as SEQ ID NO.3. It is proved through experiments that the kit can detect adverse effects of the high-pathogenicity porcine reproductive and respiratory syndrome virus (HP-PRRSV), a hog cholera virus, a C-type porcine reproductive and respiratory syndrome virus, a porcine circovirus type II, a porcine pseudorabies virus and a foot and mouth disease virus in a specificity mode. It is proved through experiments that the kit can detect out templates of at least 70 copies at the temperature of 40 DEG C on the condition of 20 min amplification, and the conformity between the kit and RT-qPCR is high. This shows that the kit can detect HP-PRRSV fast, efficiently and sensitively and provides an effective technological means for differential diagnosis of HP-PRRSV.

A process for preparing the antigen protein used for hog cholera vaccine includes extracting hog cholera virus RNA, reverse transcription to obtain the immune gene E2 of hog cholera virus, using E2 as template for PCR while inserting them to expression carrier of Bichia yeast, introducing the recombinant expression carrier to Bichia yeast, screening the recombinant Bichia yeast, discriminating its expression product testing its immunoactivity, determining the chosen recombinant yeast, testing and analyzing its culture condition, choosing the optimal culture condition, culturing the recombinant yeast and preparing the antigen protein of said vaccine.

The invention discloses a hog cholera virus truncated E2 protein which is designed on the basis of protein spatial structure, and an application of the same. In the invention, according to the crystalstructure of bovine viral diarrhea virus E2 protein, the spatial structure of the hog cholera virus E2 protein is simulated, and then the hog cholera virus E2 protein is subjected to truncated expression, wherein the amino acid sequence of the truncated protein E2B / C / D / A is represented as the SEQ ID No.1. The truncated protein can maintain the complete antigenicity of the E2 protein, and has no cross reaction with a bovine viral diarrhea virus antibody. The invention further constructs a CHO cell line which stably expresses the truncated protein E2B / C / D / A and is assigned the accession numberof CGMCC No.14722. The invention also discloses an indirect ELISA kit which is used for detection of a hog cholera virus antibody, wherein the enveloped antigen is the hog cholera virus truncated protein E2B / C / D / A. The kit is used for specifically detecting the hog cholera virus antibody with high specificity, sensitivity and repeatability.

This invention relates to a recombinant vector including a recombinant porcine adenovirus, stably incorporating and capable of expression of at least one heterologous nucleotide sequence. The nucleotide sequence is preferably one which encodes an antigenic determinant of Hog Cholera Virus or Pseudorabies virus. The further invention relates to a method of production of recombinant vectors, to methods of preparation of vaccines based on the vectors, to administration strategies and to methods of protecting pigs from disease.

The invention discloses a pair of primers and a probe for detecting hog cholera virus based on a digital PCR technology, a kit and a method thereof. The primers and the probe include an upstream primer FP, a downstream primer BP and a probe. The detection kit includes a primer set, 2*RT-ddPCR Supermix, RNase-free distilled water, a viral total RNA extraction reagent, a negative control and a positive control. The detection method comprises the following steps: extracting a sample RNA to be tested, quantitatively detecting the sample RNA to be detected by the microdrop digital PCR technique, judging whether the sample to be tested contains hog cholera virus by the copy number obtained by the amplification analysis, and determining the content thereof. The invention has the advantages of accuracy, sensitivity and wide applicability, has no dependency on a standard curve, can realize absolute quantification, can achieve early detection, early treatment and early prevention of diagnosis ofswine fever, and can effectively control the outbreak of the epidemic.

The invention relates to an enzyme-linked immuno sorbent assay (ELISA) kit created on the basis of hog cholera virus recombinant protein nopaline synthase (NS2). The ELISA kit comprises an elisa plate wrapped by the hog cholera virus recombinant protein NS2, a ten-time concentration detergent, a serum diluent, a standard serum, an anti-swine Intravenous gamma globulin (IgG) enzyme-labeled antibody, an nzyme-labeled antibody diluent, a sealing agent, a color developing agent A, a color developing agent B and a stop solution. The ELISA kit can detect hog cholera virus antibodies. The method created by the ELISA kit has specificity, sensitivity and operability. In a sensitivity test, the coincidence rate between the method of the ELISA kit and an immunofluorescence technical method is 82%, and the method of the ELISA kit is more sensitive than the immunofluorescence technical method. The detection rate of other etiological agents through the method of the ELISA kit is zero. Compared with the hemagglutination inhibition (HI) method, the method of the ELISA kit has the advantages of being fast and low in cost, and having easily judging results, and can carry out detection work after hog cholera lapinized virus vaccine immunity.

The invention relates to a cell strain of monoclonal antibodies of E2 protein resisting hog cholera virus and an application thereof. A hybridoma cell strain stably secreting the protein E2 resisting the hog cholera virus is obtained by screening, the monoclonal antibodies CSFV-1C8 secreted by the cell strain can generate specific reaction with the E2 protein of CSFV, and does not react with E2 protein of bovine viral diarrhoea / mucosal disease virus; and the monoclonal antibodies can recognize the E2 protein of the hog cholera virus specifically, and the epitope of recognition is FDFDGPDGL. The monoclonal antibodies and the epitope of the E2 protein of the hog cholera virus recognized by the monoclonal antibodies can be prepared into a reagent for detecting the CSFV, and thus laying foundation for establishing a serological detection method of antibodies of the hog cholera virus. A competitive ELISA kit for detecting the antibodies for the hog cholera and established by the invention has the advantages of specificity, sensitivity, simplicity and convenience in operation and the like and is suitable for large-scale screening of the antibodies for the hog cholera.