112 results about "Vesicular stomatitis virus VSV" patented technology

Novel purification processes for obtaining vesicular stomatitis virus (VSV) of improved purity from mammalian cell culture are described herein. More particularly, in certain embodiments, a process is described for purifying VSV from cell culture fluid of a mammalian cell culture infected with VSV, the process comprising: clarifying the cell culture fluid by low-speed centrifugation and recovering the VSV in the supernatant; filtering the supernatant through a 0.2 to 0.45 μm filter and recovering the VSV in the filtered solution; loading the VSV filtered solution onto a anion exchange membrane adsorber equilibrated with a first pH buffered salt solution, eluting the VSV from the anion exchange membrane adsorber with a second pH buffered salt solution and recovering the eluted VSV fractions; purifying the recovered VSV by tangential flow filtration (TFF) using a TFF membrane having a molecular weight cutoff between 300 kDa and 1,000 kDa and recovering the VSV in the retentate, and filtering the VSV retentate through a 0.2 to 0.22 μm filter and recovering the VSV in the filtered solution.

A pseudo-virus packaging system of coronaviruses comprises a vesicular stomatitis virus (VSV) vector with a Fluc and EGFP double reporter gene replacing a GP gene and an assembly cell expressing coronavirus spike protein S. The double reporter gene is selected from luciferase and fluorescent protein, and the luciferase reporter gene is preferably a Fluc gene. The fluorescent protein reporter geneis preferably an EGFP gene, and the assembly cell is preferably 293T. The invention further relates to a one-step packaging method of the packaging system; a pseudo-virus with single-cycle infection,low background value and high titer can be quickly packaged through simple, convenient and rapid operation, and the packaging system has the characteristic of being rapid in detection compared with aslow virus mediated pseudo-virus system; the one-step packaging method can be used for researching COVID-19, SARS, MERS and other coronaviruses and other viruses, provides a powerful screening tool for evaluation of antiviral preparations and vaccine, and has wide application value.

The present relation relates to recombinant vesicular stomatitis virus for use as prophylactic and therapeutic vaccines for infectious diseases of AIDS. The present invention encompasses the preparation and purification of immunogenic compositions which are formulated into the vaccines of the present invention.

Based on a highly conserved domain of a foot-and-mouth disease virus 3D protein coding gene, a vesicular stomatitis virus N protein coding gene and a swine vesicular disease virus VP1 protein coding gene, the invention designs a specific primer and a probe and develops a multiple fluorescence RT-PCR detection method used for simultaneously detecting the three animal viruses. The detection method can detect 102 copied plasmids containing target amplification sequences in 1 h. The method has high sensitivity and good specificity, and can achieve rapid high-throughput fluorescence RT-PCR detection for single-virus infection or mixed infection by a plurality of viruses.

The invention discloses a primer combination for identifying foot-and-mouth disease virus and vesicular stomatitis virus and application thereof. The primer combination is composed of a primer set I and a primer set II. The primer set I is composed of primers FMDV-F3, FMDV-B3, FMDV-FIP and FMDV-BIP which are sequentially disclosed as Sequence 1-4. The primer set II is composed of primers VSV-F3, VSV-B3, VSV-FIP and VSV-BIP which are sequentially disclosed as Sequence 5-8. The invention also discloses application of the primer combination in identifying foot-and-mouth disease virus and vesicular stomatitis virus, application in identifying whether a virus to be detected is foot-and-mouth disease virus or vesicular stomatitis virus, and application in identifying whether a sample to be detected is infected by foot-and-mouth disease virus and / or vesicular stomatitis virus. The duplex RT-LAMP (reverse transcription-loop-mediated isothermal amplification) method established by the invention is a simple quick low-cost diagnosis method, can be used for grass-root and field quarantine inspection under poor conditions, and is also suitable for large-scale epidemiological survey.

The invention relates to a novel gene sequence of a recombinant chicken Alpha interferon, constructs a recombinant expression vector thereof and belongs to a gene engineering biological product obtained by a molecular biology method. The gene sequence of a newly designed chicken Alpha interferon is recombined into a pPICZ Alpha-A vector and then is confirmed on the special position of a microzyme by an electric conversion mode. Besides, a pichia expression system is adopted to express a foreign gene, thus being beneficial to the commercial production of the chicken interferon. The protein expressed by a gene group after being diluted by 4365158.3 times can completely restrain the attraction of vesicular stomatitis virus of 100-1000TCID50. Compared with a natural chicken Alpha interferon, the novel gene sequence of a recombinant chicken Alpha interferon has a higher anti-virus effect; the anti-virus effect thereof is improved by 8 times. Test also detects that the protein expressed by the gene can restrain the proliferation of a newcastle disease virus and an avian influenza virus; besides, the effect of the interferon with high concentration is more remarkable.

The invention belongs to the technical field of bovine virus detection and particularly relates to a foot and mouth disease virus and vesicular stomatitis virus identifying duplex fluorescence RT-LAMP (loop-mediated isothermal amplification) detection primer group, a kit and application thereof. The foot and mouth disease virus and vesicular stomatitis virus identifying duplex fluorescence RT-LAMP detection primer group comprises two groups of specific primers, wherein one group is FMDV-F3, FMDV-B3, FMDV-FIP (F1c-F2) and FMDV-BIP (B1c-B2), and the other group is VSV-F3, VSV-B3, VSV-FIP (F1c-F2) and VSV-BIP (B1c-B2), and sequences of the two groups are shown as SEQ ID NO.1 and SEQ ID NO.8 respectively. An established foot and mouth disease virus and vesicular stomatitis virus identifying duplex fluorescence RT-LAMP method has advantages of simplicity, convenience, quickness, specificity, sensitivity and the like and can be used for FMDV (foot and mouth disease virus) and VSV (vesicular stomatitis virus) clinical detection and epidemiological investigation. The FMDV and VSV duplex fluorescence RT-LAMP method is a simple, quick and low-cost diagnosis method and suitable for large-scale epidemiological investigation.

The invention discloses a gene chip and a detection method for detecting FMDV, VSV, SVDV, PPRV and BTV. The detection method comprises the step of detecting foot and mouth disease viruses (type A, Asian type I and type O), vesicular stomatitis virus, swine vesicular disease virus, Peste des petits ruminants virus and bluetongue virus. The method comprises the following specific steps: designing a PCR primer by virtue of sequence analysis of standard strain genome, and performing cloning and sequence analysis on target genes; designing a specific probe, and simultaneously detecting the foot and mouth disease viruses, vesicular stomatitis virus, swine vesicular disease virus, Peste des petits ruminants virus and bluetongue virus. The invention aims at establishing a method for detecting the foot and mouth disease viruses, vesicular stomatitis virus, swine vesicular disease virus, Peste des petits ruminants virus and bluetongue virus by adopting a microarray chip which is high in sensitivity and high in specificity and through which the time and labor are saved and the result is easily observed.

The invention discloses a primer combination for simultaneously identifying 8 kinds of cattle pathogens and a GeXP detection method. The primer combination provided by the invention consists of a primer pair I, a primer pair II, a primer pair III, a primer pair IV, a primer pair V, a primer pair VI, a primer pair VII and a primer pair VIII. The invention also provides the GeXP detection method for simultaneously identifying foot and mouth disease viruses, blue tongue viruses, vesicular stomatitis viruses, bovine viral diarrhoea viruses, bovine rotaviruses, enterotoxigenic escherichia coli, infectious bovine rhinotracheitis viruses and peste des petits ruminants viruses. The GeXP detection method built by the invention can be used for simultaneously identifying 8 kinds of cattle infectious disease pathogens. The method has the characteristics of high flux, specificity and high sensitivity, and can be used for the cattle disease epidemiology monitoring and the emergent epidemic situation identification and diagnosis, and guarantees the healthy development of cattle rearing industry.

The invention belongs to the technical field of biological genetic engineering, and discloses technology for constructing and expressing porcine alpha interferon (PoIFN-alpha) and interleukin 2 (PoIL-2) chimeric gene and quickly renaturing and purifying expression protein. The technology constructs mature peptide genes of PoIFN-alpha and PoIL-2 into PoIFN-alpha-linker-PoIL-2 chimeric gene through a genetic flexible linker (linker) (G4S)3 and clones the chimeric gene into a pGEM-T Easy vector by adopting an overlap extension PCR method, and subclones the chimeric gene into a pQE-30 expression vector for prokaryotic expression. The recombinant fusion protein (rPoIFN-alpha-linker-PoIL-2) can be quickly renatured and purified through urea modification, renaturation by low concentration protein renaturing solution, PBS solution dialysis and other steps. The purified rPoIFN-alpha-linker-PoIL-2 fusion protein has proliferation activity for inhibiting vesicular stomatitis virus (VSV) on a cell, is used for prevention and treatment of porcine virosis as a main ingredient of an antivirus preparation, and has high efficiency, broad spectrum, safety and low price.

The invention discloses a recombinant human hepatitis B virus core protein fused protein. The fused protein comprises a protein (X), a linker peptide (L) and a hepatitis B virus core protein (HBc) from the end N to the end C in sequence; the linker peptide (L) has the amino acid sequence of Gly-Ser-(Gly-Gly-Gly-Gly-Ser)n, and n is an integer between 2 and 20 and is 9 or 18, particularly; the end C of the linker peptide (L) is connected with the end N of the hepatitis B virus core protein (HBc); the end C of the protein (X) is connected with the end N of the linker peptide (L); and the protein (X) is a red fluorescent protein or vesicular stomatitis virus G glycoprotein. The hepatitis B virus core protein (HBc) is connected with the functional protein, and the functions of the proteins on two ends of the linker peptide (L) can be both guranteed. The problem in the prior art that the functions of the hepatitis B virus core protein (HBc) and the functions of the functional protein can not be both guaranteed after the hepatitis B virus core protein (HBc) is fused with the functional protein is solved. The fused protein is of great importance to the research on the hepatitis B virus (HBV).

The invention discloses a recombinant porcine alpha interferon, prepared by the following methods: A1, synthesizing an artificially modified porcine alpha interferon; A2, constructing a recombinant eukaryotic expression vector pGAPZ alpha-IFN alpha; and A3, highly expressing IFN alpha by an eukaryotic cell yeast expression system. The invention is characterized by taking a supernatant to purifying the porcine alpha interferon which is cultured by a lot of engineering bacteria and expressed with a chromatography column, then collecting a target protein eluate, filtering through a 0.22 mum microfiltration membrane, then determining the activity by cytopathic inhibition, and calculating the potency unit according to 50% pathology; wherein the cell used for determination is Madin-Darby bovine kidney (MDBK), and vesicular stomatitis virus (VSV) is used for attacking the virus. According to the invention, the recombinant porcine alpha interferon obtained by purification is applied in controlling PCMV, and experiments prove that the recombinant porcine alpha interferon has obvious protection effect on PCMV infected cells. The route of administration of the porcine alpha interferon is injection or mucous membrane administration, and the dosage form comprises injection or nasal drops.

The invention relates to a coronavirus pseudovirus packaging system. The coronavirus pseudovirus packaging system comprises a vesicular stomatitis virus VSV vector and an assembly cell, wherein the vesicular stomatitis virus VSV vector is formed by replacing GP genes with Fluc and EGFP double reporter genes, and the assembly cell is used for expressing coronavirus spike protein S. The double reporter genes are selected from luciferase and fluorescent protein, and the luciferase reporter gene is preferably the Fluc gene. According to the packaging system, a one-step packaging method is adopted, so that pseudoviruses which are infected in a single cycle, low in background value and high in titer and have the characteristic of rapid detection compared with a lentivirus-mediated pseudovirus system can be rapidly packaged, and the packaging system can be used for researching coronaviruses such as COVID-19 (SARS-CoV-2), SARS (SARS-CoV) and MERS; and the pseudoviruses can be used for evaluating the efficacy of a disinfectant through the steps of a virus pollution distribution model, scene building and sampling detection, a safe, convenient and effective tool method is provided for evaluating the disinfectant, and the pseudoviruses have wide application value.

The present invention provides for pharmaceutical compositions comprising pancreatic enzyme preparations (PEPs) with viral infectivity reduced below significant levels and having high enzymatic activity. The PEPs can comprise lipases, proteases, amylases, non-enveloped viruses (e.g., porcine parvovirus (PPV), porcine circovirus type 2 (PCV-2), porcine encephalomyocarditis virus (EMCV)), and enveloped viruses (e.g., vesicular stomatitis virus (VSV), and influenza A (IFA)). The present invention also includes methods of treating pancreatic insufficiency by administering these pharmaceutical compositions and methods of making the same by treating the PEP with beta-propiolactone (BPL) to reduce viral infectivity.

The invention discloses a vesicular stomatitis virus vector-based novel coronavirus chimeric recombinant vaccine as well as a preparation method and application thereof. The active component of the recombinant vaccine is recombinant virus rVSV-SARS-CoV / 2-RBD, and is a virus obtained by replacing glycoprotein G of the vesicular stomatitis virus with chimeric envelope protein S; the chimeric envelope protein S is a protein obtained by replacing the RBD of the SARS-CoV envelope protein S with the RBD of the SARS-CoV-2 envelope protein S; the amino acid sequence of the RBD of the SARS-CoV envelopeprotein S is the 315-536 site of the amino acid sequence of the SARS-CoV envelope protein S; and the amino acid sequence of the RBD of the SARS-CoV-2 envelope protein S is the 319-541 site of the amino acid sequence of the SARS-CoV-2 envelope protein S. The recombinant virus has great significance to vaccine development of new coronaviruses.

The invention discloses canine recombinant interferon alpha, and also provides a preparation method of the canine recombinant interferon alpha. Recombinant genes of canine interferon alpha are expressed in escherichia coli, and canine interferon alpha with a purity of more than 98% is obtained through processes of renaturation and purification. The biological activity is determined by a MDCK (canine kidney cell)-VSV (vesicular stomatitis virus) virus system, and the specific activity reaches 2-3*107 IU / mg.

The present invention is directed to a method of reducing the viability of a tumor cell involving administering a virus that is not a common human pathogen to the tumor cell. Preferably, the virus exhibits differential susceptibility, in that normal cells are not affected by the virus. This differential susceptibility is more pronounced in the presence of interferon. The tumor cell is characterised by having low levels, or no PKR activity, or as being PKR- / -, STAT1- / - or both PKR- / - and STAT- / -. The virus is selected from the group consisting of Rhabdovirus and picornavirus, and preferably is vesicular stomatitis virus (VSV) or a derivative thereof.

The invention provides a preparation method and application of 4-1BB-containing lentivirus. According to the lentivirus prepared by the preparation method for the lentivirus, CD28 can be not required to be added; only a small amount of CD3 is directly added to a culture medium to serve as a first T-cell activating signal before transfection, so that a process of preparing Car-T cells is reduced, and the cost and pollution risk are reduced; T-cells are stimulated to be further activated through recombinant VSVG (Vesicular Stomatitis Virus Glycoprotein), so that the contact probability of a virus and cells is improved to effectively improve infection efficiency.

The invention discloses a Siberian tiger alpha interferon and a coding gene and application thereof. A Siberian tiger alpha interferon family is obtained by cloning a Siberian tiger genome, an amino acid sequence of its mature peptide is shown as SEQ ID No.1-11, and the nucleotide sequence of the coding gene is shown as SEQ ID No.12-22. A Siberian tiger IFN-alpha family protein has a shared conservative structure domain, namely an IFabd structure domain. Antiviral activity functional verification results prove that the Siberian tiger IFN-alpha family protein has the functions of resisting vesicular stomatitis virus, canine distemper and bird flu virus. The invention further discloses an antiviral drug composition, including an effective dose of the Siberian tiger alpha interferon for prevention or treatment and pharmaceutically acceptable excipients or carriers. A solid theoretical basis is laid for the Siberian tiger alpha interferon for the further study of other viral diseases and appearing of Siberian tiger alpha interferon products.

The invention provides a porcine interferon alpha and an application thereof. The porcine interferon alpha is synthesized through selecting high-expression codon design according to the codon bias of the of a Escherichia coli prokaryotic expression system, the amino acid sequence of the porcine interferon alpha is represented by SEQ ID NO. 1, and the nucleotide sequence for encoding the porcine interferon alpha is represented by SEQ ID NO. 2. A method for preparing the porcine interferon alpha has the advantages of simplicity, low cost, easiness in industrial production, high expression level of the porcine interferon alpha, thorough denaturation in the inclusion body treatment process, and high protein renaturation rate porcine. The porcine interferon alpha prepared in the invention has high titer, can inhibit 100TCID50 vesicular stomatitis virus, has a specific activity reaching up to 10<8>, has good bioactive functions, can be used to prepare medicines for preventing and treating the swine virus infection and enhancing the immune function, and has a broad market application prospect.

The invention relates to a kit for detecting vesicular stomatitis antibody and a preparation method thereof. The kit for detecting vesicular stomatitis antibody comprises a reagent I and a reagent II placed respectively, wherein the reagent I is a vesicular stomatitis virus antihelion peridium enzyme linked immunosorbent assay plate and the reagent II is monoclonal antibody IgG-HRP enzyme conjugates, wherein the monoclonal antibody IgG-HRP enzyme conjugates is idiocrasy monoclonal antibody of mouse-anti vesicular stomatitis virus of horse radish peroxidase mark. The invention also provides a preparation method of the kit. The kit adopts competitive ELISA to detect the vesicular stomatitis in serum, and can be used for rapid diagnosis, detection, quarantine and epidemiological investigation of vesicular stomatitis.

A process is described for purifying vesicular stomatitis virus (VSV) from cell culture fluid of a mammalian cell culture infected with VSV, the process comprising: clarifying the cell culture fluid by low-speed centrifugation and recovering the VSV in the supernatant; filtering the supernatant through a 0.2 to 0.45 μm filter and recovering the VSV in the filtered solution; loading the VSV filtered solution onto a anion exchange membrane adsorber equilibrated with a first pH buffered salt solution, eluting the VSV from the anion exchange membrane adsorber with a second pH buffered salt solution and recovering the eluted VSV fractions; purifying the recovered VSV by tangential flow filtration (TFF) using a TFF membrane having a molecular weight cutoff between 300 kDa and 1,000 kDa and recovering the VSV in the retentate, and filtering the VSV retentate through a 0.2 to 0.22 μm filter and recovering the VSV in the filtered solution.

The invention belongs to the field of biotechnology, and particularly relates to an oncolytic virus vaccine and a tumor treatment drug combing oncolytic virus vaccine with an immune checkpoint inhibitor. A brand-new oncolytic virus attenuated strain is provided by site-directed mutagenesis of wild-type virus matrix protein M of VSV (vesicular stomatitis virus). The attenuated strain can be used asa drug alone to treat tumors, and has safety and cure rate higher than those of wild-type viruses and other known attenuated strains. On the basis of the oncolytic virus attenuated strain, a vaccinethat can be used in tumor treatment is also provided by inserting NY-ESO-1 into the attenuated strain. The vaccine has high cure rate and high biological safety. On the basis of the vaccine, the invention also combines the vaccine with the immune checkpoint inhibitor to provide the drug capable of efficiently treating various tumors. In mouse lung cancer models, the cure rate can be astonishing, up to 87.5%, and the treatment effect on large tumors is also good.

The present invention discloses an oral avian interferon fusion protein and applications of the oral avian interferon fusion protein as an immunopotentiator, wherein the fusion protein comprises chicken alpha interferon and chicken beta interferon. The invention further provides applications of the fusion protein, wherein the applications comprise the application of the fusion protein as an immunopotentiator, the application of the fusion protein as an oral immunopotentiator, the preparation of an immunopotentiator, the preparation of an oral immunopotentiator, the application of the fusion protein as a vaccine immunopotentiator, the application of the fusion protein as an oral vaccine immunopotentiator, the application of the fusion protein as an avian influenza virus vaccine immunopotentiator, the application of the fusion protein as an oral avian influenza virus vaccine immunopotentiator, the preparation of a vaccine, the preparation of an avian influenza virus vaccine, the application of the fusion protein as an anti-virus preparation, the preparation of an anti-virus preparation, the application of the fusion protein as an anti-vesicular stomatitis virus preparation, and the preparation of an anti-vesicular stomatitis virus preparation. According to the present invention, the fusion protein as the immunopotentiator or anti-virus preparation can be subjected to oral administration, and provides good clinical effects for the improvement of the early immunity of chickens.

The invention discloses a primer combination and GeXP detection method for simultaneously identifying 5 bovine viral dermatitis viruses. The primer combination is composed of a primer pair I, a primer pair II, a primer pair III, a primer pair IV and a primer pair V. The invention also discloses a GeXP detection method for simultaneously identifying foot-and-mouth disease virus, bluetongue virus, vesicular stomatitis virus, bovine viral diarrhoea virus and infectious bovine rhinotracheitis virus. The GeXP detection method can simultaneously identify the 5 bovine viral dermatitis viruses. The method has the characteristics of higher flux, higher specificity and higher sensitivity, can be used for monitoring of bovine disease epidemiology and differential diagnosis of unexpected epidemic situations, and ensures the healthy development of cattle raising industry.

The invention discloses a novel recombinant coronavirus based on a vesicular stomatitis virus vector as well as a preparation method and application of the novel recombinant coronavirus. The novel recombinant coronavirus is a virus obtained by replacing glycoprotein G of vesicular stomatitis virus with envelope protein S; the envelope protein S comprises an envelope protein S extracellular region of 2019-nCoV (SAR-CoV-2) or a part of the sequence of the envelope protein S extracellular region. Experiments prove that the recombinant novel coronavirus can simulate the process that 2019-nCoV (SARS-CoV-2) invades cells, and can stimulate an organism to generate immunoreaction aiming at the 2019-nCoV (SARS-CoV-2), so that the recombinant novel coronavirus is an ideal tool for researching the infection process of the 2019-nCoV (SARS-CoV-2), can be developed into vaccines, and has a good application prospect. the recombinant novel coronavirus has important significance for screening inhibitors for inhibiting virus invasion and developing vaccines.

The present invention relates to an immunogenic or vaccine composition to induce an immune response or protective immune response against vesicular stomatitis virus (VSV) in an animal susceptible to VSV. The composition may include a pharmaceutically or veterinarily acceptable vehicle or excipient, and a vector. The vector may contain at least one heterologous nucleic acid molecule(s), expresses in vivo in the animal VSV antigen(s), immunogen(s) or epitope(s) thereof, e.g., VSV G protein and / or VSV N protein and / or VSV M protein. The heterologous nucleic acid molecule(s) may be adjusted to the vector / mammalian cell system by codon optimization. The composition can contain an adjuvant, such as carbomer. Methods for making and using such a composition, including prime-boost regimes and including as to differential diagnosis, are also contemplated.

The invention discloses a gene chip and kit used for detecting foot and mouth disease viruses, vesicular stomatitis viruses and swine vesicular disease viruses. The gene chip and kit and a detection method provided by the invention can accurately and effectively detect foot and mouth disease viruses, vesicular stomatitis viruses and swine vesicular disease viruses and have the advantages of high specificity and sensitivity, short detection time, rapid detection and good application prospects.

The present invention provides an antimicrobial composition, which comprises a lauroyl lauroyl arginate (LAE) ion pair derivative, and the antimicrobial composition is an antiviral agent against an enveloped virus, wherein the enveloped virus is selected from vesicular stomatitis virus of the herpes virus, or the Newcastle disease virus selected from the paramyxovirus. The antiviral agent is the anti-infective agent comprising a condensate derived from fatty acid and esterified binary amino acid, and is capable of acting on Newcastle disease virus and vesicular stomatitis virus in-vitro, and reducing the infection of Newcastle disease virus and vesicular stomatitis virus to cells. The antiviral agent is added to the cultured Newcastle disease virus (NDV) and vesicular stomatitis virus (VSV), or the antiviral agent is added at different times before the virus is infected and after the infection, the infected cells are detected, and the number of viruses is found to be significantly reduced. The antiviral agent can effectively inhibit or kill the virus and can be used to prevent or reduce the ability of the virus to infect cells. The present invention also provides a method of preparing the antimicrobial composition, which has the characteristics of better virucidal effect and lower dosage than that of the original antimicrobial composition containing a LAE compound.