48 results about "Unstructured Proteins" patented technology

The invention discloses an mRNA (messenger ribonucleic acid) transcription skeleton vector and application thereof in preparation of a self-amplification type mRNA vaccine, and belongs to the technical field of veterinary biological products. According to the invention, alphavirus non-structural protein regions NSP2 and NSP3 of a self-amplification type mRNA skeleton are modified, a polyA tail is optimized, and a novel self-amplification type mRNA skeleton is constructed; the self-amplification type mRNA vaccine can improve the antigen expression efficiency, reduce the toxicity of non-structural protein to cells and improve the encapsulation efficiency and delivery efficiency by combining with the optimization of the components and proportion of a lipid nano delivery system, and meanwhile, the self-amplification type mRNA vaccine is wide in applicable pathogen antigen range and can be popularized to more animal epidemic diseases. The characteristics of high-efficiency expression, low-toxicity delivery and single-dose immunization are expected to renovate the immunization strategy of the existing animal vaccine, the epidemic prevention cost of the breeding industry is reduced, and the method has a wide industrial application prospect.

This invention provides a dengue / Zika virus genetically engineered vaccine and its application. The dengue / Zika virus vaccine comprises an open reading frame encoding envelope protein domain III (EDIII) and the non-structural protein NS1, and displays EDIII monomers in the delivery vector shell. Immunization with this vaccine primarily induces a type-specific antibody response, reducing the production of cross-antibodies and thus effectively avoiding or eliminating the risk of antibody-dependent enhancement of infection (ADE). This vaccine can be used to prevent dengue virus and Zika virus infection.

A lipid nanoparticle composition for use in the delivery of a self-amplifying RNA (saRNA) construct in one or more target cells is disclosed. The lipid nanoparticle composition comprises a lipid mixture comprising at least one (ionizable) cationic lipid, at least one helper lipid, a sterol, a corticosteroid such as Dexamethasone, and a least one lipid-polyethylene glycol conjugate. The saRNA construct comprises a first open reading frame which encodes one or more non-structural proteins, and a second open reading frame operatively linked to the first open reading frame. The second open reading frame comprises a coding region which encodes one or more target proteins. In some embodiments, the target proteins comprise one or more therapeutic proteins. In some embodiments, the target proteins comprise one or more one or more Cas proteins and / or variants thereof for use in CRISPR-based gene editing.

This invention belongs to the field of genetic engineering. By introducing a TC tag between amino acids 199 and 200 of the wild-type bluetongue virus non-structural protein NS2, a recombinant virus was rescued. Analysis of viral plaques and growth curves revealed that the recombinant virus formed plaques of similar size to the wild-type virus, and their growth curves showed no significant difference. Immunofluorescence and FlasH-EDT2 staining of NS2 showed that the fluorescence of NS2 labeled with anti-NS2 antibody overlapped with that labeled with FlasH-EDT2. Three-dimensional imaging of FlasH-EDT2-labeled NS2 revealed that the viral inclusion bodies exhibited fluorescence around the periphery but no fluorescence inside, displaying an overall "core-shell" hierarchical structure. This recombinant bluetongue virus can be used for tracing and localizing NS2 protein and viral inclusion bodies in living cells, and can also be applied to BTV neutralization assays, antiviral drug screening, and other related research.

The invention provides a norovirus genome splicing method and device, a medium and a product. The method comprises the following steps: acquiring a norovirus gene database, wherein the norovirus gene database comprises a non-structural protein coding region sub-library and a capsid protein coding region sub-library; obtaining original sequencing data; performing quality control on the original sequencing data to obtain screened sequencing data; comparing and assembling the screened sequencing data with the non-structural protein coding region sub-library, and determining at least one first recombinant coding region; and comparing and assembling the screened sequencing data with the capsid protein coding region sub-library, and determining at least one second recombinant coding region, and splicing the at least one first recombinant coding region and the at least one second recombinant coding region to obtain a recombinant genome sequence. Through the technical scheme disclosed by the invention, efficient and accurate splicing of the recombinant norovirus genome sequence is realized, and the assembly accuracy and efficiency of the recombinant norovirus genome sequence are improved.

The application discloses a recombinant FCV antigen and a construction method and application thereof. The construction method of the recombinant FCV antigen comprises the following steps: fusing a T cell epitope coding sequence of a non-structural protein NS7 of FCV and a coding sequence of a SpyTag peptide segment through a coding sequence of a linker, then cloning into a baculovirus transfer vector to obtain a recombinant plasmid, and finally integrating the T cell epitope coding sequence into Bacmid through Tn7 transposition to finally obtain the recombinant FCV antigen. The application selects NS7 as a core immunogen, guides the immune system to produce a high cellular immune response, and thus makes up for the deficiency of an existing vaccine in clearing intracellular viruses; meanwhile, a specific T cell epitope is selected in the sequence of NS7 as an immunogen, which can avoid the immunological escape caused by the variation degree of antigens among different strains and virus antigen drift, and thus provides broader protection.

The present application relates to a replicable RNA molecule and the use thereof. The replicable RNA molecule comprises, from the 5′ end to the 3′ end, a 5′ cap, a 5′ UTR, an open reading frame encoding an RNA replicase, a promoter, a sequence of interest, a 3′ UTR and a poly(A) tail, wherein the RNA replicase is capable of amplifying the replicable RNA molecule and is capable of amplifying an RNA molecule containing the sequence of interest and 3′ UTR, wherein the RNA replicase is a nonstructural protein or a functional variant thereof derived from Mosso das Pedras virus (MDPV), Everglades virus (EVEV), Rio Negro virus (RNV), Mucambo virus (MUCV), Highlands J virus (HJV), Pixuna virus (PIXV), Trocara virus (TROV), Cabassou virus (CABV), Tonate virus (TONV), Bebaru virus (BEBV), Fort Morgan virus (FMV), Getah virus (GETV) or Ndumu virus (NDUV).

Synthetic peptides mimicking the nsp10 sequence in the region interacting with nsp16 capable of penetrating cell membranes and inhibiting SARS-CoV-2 replication for the treatment of moderate to severe COVID-19. The invention relates to peptides inhibiting SARS-CoV-2 replication, likely through inhibition of Methyltransferase complexes (NSP10 / NSP16 and NSP10 / NSP14). The peptide of the present invention, P3, contains sequences corresponding to amino acids 89-96 of the non-structural protein 10 (NSP10) of SARS-CoV-2, with the only Cysteine modified to a Methionine. This peptide was made based on two previous designs P1 and P2, which constituted the amino acids 68-96 of the NSP10 protein of SARS-CoV-2.

The invention discloses a protein degradation product and application of non-structural protein NSs of rift valley fever virus or a coding gene of the non-structural protein NSs in preparation of the protein degradation product, and relates to the technical field of protein degradation. On the basis of the property that NSs and recombinant protein of the NSs can autonomously form a fiber-shaped high polymer structure, a degradation machine capable of degrading target protein is self-assembled, so that the specific and efficient degradation of the target protein is realized. Therefore, the new application of the non-structural protein NSs of the rift valley fever virus in targeted degradation of the protein is found, specific efficient degradation of the disease-related protein can be achieved, and the effect of treating diseases is expected to be achieved.

The present disclosure provides novel self-amplifying RNA (saRNA) constructs that demonstrate enhanced protein expression, prolonged durability, reduced immunogenicity, and the ability to express multiple therapeutic proteins homogeneously. The saRNA constructs comprise a 5' untranslated region (5'UTR), non-structural protein genes derived from alphaviruses, at least one gene of interest encoding a therapeutic protein, a 3' untranslated region (3'UTR), and one or more modified nucleosides. Also disclosed are dual construct systems comprising a first construct encoding non-structural proteins and a second construct encoding one or more genes of interest. Methods of producing and using the saRNA constructs for engineering cells, particularly immune cells, for treatment of various conditions including cancer, inflammatory conditions, and infectious diseases are provided. The saRNA constructs enable the generation of "armored" immune cells expressing multiple therapeutic proteins, thereby providing a multi-pronged approach to complex diseases.

This invention discloses a method for preparing pet stem cell exosomes, belonging to the field of animal medicine technology. This invention treats stem cells with an inducing agent to secrete exosomes with high TFR expression on their surface. These exosomes specifically target the conserved region of the NS1 gene, a non-structural protein of the parvovirus family, allowing the internally loaded siRNA and trefoil factor 3 to target and enter infected cells, directly inhibiting viral replication at the gene level to treat feline panleukopenia virus (FPV) and canine parvovirus infections and alleviate sequelae. This invention solves the problems of existing technologies, such as difficulty in targeting, lack of specific drugs, reliance on intensive supportive care leading to a short treatment window, inability to directly clear the virus, risk of cross-infection, and potential long-term sequelae. It has the potential to be applied to the preparation of drugs for treating canine parvovirus and feline panleukopenia.

The present disclosure provides novel self-amplifying RNA (saRNA) constructs that demonstrate enhanced protein expression, prolonged durability, reduced immunogenicity, and the ability to express multiple therapeutic proteins homogeneously. The saRNA constructs comprise a 5' untranslated region (5'UTR), non-structural protein genes derived from alphaviruses, at least one gene of interest encoding a therapeutic protein, a 3' untranslated region (3'UTR), and one or more modified nucleosides. Also disclosed are dual construct systems comprising a first construct encoding non-structural proteins and a second construct encoding one or more genes of interest. Methods of producing and using the saRNA constructs for engineering cells, particularly immune cells, for treatment of various conditions including cancer, inflammatory conditions, and infectious diseases are provided. The saRNA constructs enable the generation of "armored" immune cells expressing multiple therapeutic proteins, thereby providing a multi-pronged approach to complex diseases.

The invention discloses a human-sheep rotavirus gene reassortment attenuated strain LG1 and application of the human-sheep rotavirus gene reassortment attenuated strain LG1. The human-sheep rotavirus gene reassortment attenuated strain LG1 comprises: i) a coding gene of a structural protein VP7 of a human G1 type rotavirus HN15D2 strain; and ii) coding genes of structural proteins VP1, VP2, VP3, VP4 and VP6 and non-structural proteins NSP1, NSP2, NSP3, NSP4 and NSP5 of the sheep rotavirus strain LLR. The human-sheep rotavirus gene reassortment strain which takes a sheep rotavirus strain as a skeleton, contains a human G1-VP7 gene and can be stably passaged on a Vero cell is successfully constructed for the first time, and the reassortment strain not only has the safety of an animal strain, but also has the immunogenicity of a human G1 type RV, can effectively prevent infection of a human G1 type wild strain, and has a good application prospect. The method has important significance for developing multivalent RV vaccines containing G1 genotypes and developing RV vaccines which are higher in prevention pertinence and provide a better protection effect on Chinese infantile diarrhea.

The application provides application of rotavirus non-structural protein 4 in improving immune function of recombinant rotavirus subunit vaccine, wherein the application is to express the rotavirus non-structural protein 4 gene and rotavirus antigen gene in different surface rings on a norovirus P particle to obtain a recombinant protein, and the recombinant protein is the recombinant rotavirus subunit vaccine. The application improves the immune function of the recombinant rotavirus subunit vaccine by using the NSP4 protein, and parallel expression of the NSP4 gene and the VP8 antigen gene or the NSP4 antigen gene can improve the NSP4 antigen and the VP8 antigen specific IgG antibody titer, improve the neutralization ability of the VP8 antigen to the rotavirus, and improve the protection of the NSP4 antigen and the VP8 antigen to the diarrhea mice induced by the rotavirus SA11 strain.

The invention provides preparation and application of an adenovirus non-structural protein, and aims at a novel chimpanzee adenovirus to screen the non-structural protein capable of detecting the titer of the chimpanzee adenovirus with high sensitivity and strong specificity. Immunofluorescence detection is carried out on the virus titer through the antibody prepared from the non-structural protein, the result is close to the detection result of a fluorescence counting method of flow cytometry, compared with detection of hypervariable region protein antibodies and whole virus antibodies in existing structural protein, the accuracy and sensitivity of detection are remarkably improved, very high specificity can be kept, and the method is suitable for large-scale popularization and application. Therefore, the chimpanzee adenovirus titer can be detected more simply, conveniently and efficiently.

The invention discloses a recombinant FCV antigen and a construction method and application thereof. The construction method of the recombinant FCV antigen comprises the following steps: fusing a T cell epitope coding sequence derived from a non-structural protein NS7 of FCV with a coding sequence of a SpyTag peptide fragment through a coding sequence of a linker, and cloning to a baculovirus transfer vector to obtain a recombinant plasmid; and finally, integrating the T cell epitope coding sequence into Bacmid through Tn7 transposition to finally obtain the recombinant FCV antigen. According to the invention, NS7 is selected as a core immunogen to guide an immune system to generate high cellular immune response, so that the defect of an existing vaccine in removing viruses in cells is overcome; meanwhile, T cell epitopes with specificity are selected from the sequence of NS7 as immunogens, so that the variability of antigens among different strains and immune escape caused by virus antigen drift can be avoided, and wider-spectrum protection is provided.

The invention discloses a rotavirus reassortment attenuated strain LG12 and application of the rotavirus reassortment attenuated strain LG12. The rotavirus reassortment attenuated strain LG12 comprises: i) a coding gene of a structural protein VP7 of a human G12 type rotavirus strain G12P [6]; and ii) coding genes of structural proteins VP1, VP2, VP3, VP4 and VP6 and non-structural proteins NSP1, NSP2, NSP3, NSP4 and NSP5 of the sheep rotavirus strain LLR. The reassortment strain provided by the invention has the safety of animal strains and the immunogenicity of human G12 type RV, can effectively prevent infection of human G12 type wild strains, and can have different degrees of cross protection on infection of other various G type RV wild strains; the method has great significance for developing RV vaccines which cover more G and P genotypes of Chinese epidemic RVA, have stronger prevention pertinence and provide a better protection effect on Chinese infantile diarrhea.

The application discloses an epitope tandem polypeptide of porcine reproductive and respiratory syndrome and related biological materials and application thereof, and belongs to the field of viral antigen vaccines or biomedical technologies. The application provides an epitope tandem polypeptide which is composed of neutralizing antigen epitopes, Th epitopes and CTL epitopes of multiple structural proteins and non-structural proteins of PRRSV in series, and includes PNB, PTH and PCTL. The epitope tandem polypeptide provided by the application can be directly used for preparing a subunit vaccine of PRRSV, and has the advantages of simple preparation method, strong immunogenicity and good safety. The epitope tandem polypeptide can also be combined with an inactivated vaccine of PRRSV and applied to enhance cellular immunity induced by the inactivated vaccine, so that the epitope tandem polypeptide can more effectively prevent and protect pigs from PRRSV infection. The epitope tandem polypeptide composition contains common epitope sequences of seven representative strains (LV, VR-2332, CH-1a, JXA1, NADC30, NADC34 and RFLP 1-4-4) of PRRSV, and is expected to have good cross-protection for infection of different strains.

The application discloses a recombinant novel goose parvovirus infectious clone plasmid and application, the infectious clone plasmid includes the full-length genome of novel goose parvovirus, and a synonymous mutation marker is introduced at the cysteine codon position in the non-structural protein coding region, under the condition that the amino acid sequence is not changed and the offspring virus replication and infection ability are not influenced, a stable single nucleotide genetic marker is provided for the novel goose parvovirus.The infectious clone plasmid can be used for rescuing the recombinant virus carrying a specific synonymous mutation marker, the obtained recombinant virus has good duck embryo adaptability and cell infection ability, and is suitable for application scenes such as cell infection and immunofluorescence detection.The application provides an important technical tool for novel goose parvovirus genetic marker tracking, pathogenic mechanism research, genetic variation analysis and vaccine research and development.

The identified peptide sequences form moderate to strong heterodimeric complexes with SARS-COV2 (B.1.617.2) virus Spike envelope proteins and a range of virus nonstructural protein sequences NSP1-14. These identified peptides are proposed to use in antiviral immunotherapy and supportive combination with chemical compound(s) for chemotherapy. The sequence ID of the identified peptides are, Seq1: ISD1, Seq2: ISD2, Seq3: ISD3, Seq4: ISD4, Seq5: ISD5, Seq6: ISD6, Seq7 Ip2, Seq8: Ip1, Seq9: Ip3, Seq10: Ip5, Seq11: Ip7, Seq12: Ip8, Seq13: Ip9, Seq14: SOI1, Seq15: SOI2, Seq16: SOI5, Seq17: SOI7, Seq18: SOI9, Seq19: SO10, Seq20: SO11.

The invention belongs to the technical field of virus construction, and particularly relates to a bovine parainfluenza virus type 3 V protein deletion strain and application thereof. The construction process of the strain comprises the following steps: by taking pBPIV3-SX full-length infectious clone plasmid as a template, respectively amplifying two gene segments before and after a base editing site of a V gene, and fusing the obtained amplified segments to obtain a segment containing V gene deletion; and replacing a fragment containing V gene deletion to BssHII and PacI restriction enzyme cutting sites of the pBPIV3-SX plasmid, and transfecting to carry out virus rescue, thereby obtaining the recombinant plasmid pBPIV3-SX. According to the invention, on the basis of a reverse genetics operation technology, a V protein deleted infectious clone plasmid is constructed, and a recombinant deleted virus is successfully rescued; meanwhile, the V protein is obtained through an escherichia coli expression system, a mouse is immunized to obtain a polyclonal antibody, a theoretical basis is provided for functional research of the BPIV3 non-structural V protein and development of genetic engineering vaccines, and the BPIV3 non-structural V protein has important scientific value and application value.

The invention relates to a novel anti-dengue virus (DENV) monoclonal antibody, a preparation method of the antibody, and related application of the antibody in immunodetection. The anti-DENV monoclonal antibody is targeted to a non-structural protein 1 (NS1) of DENV, and shows high sensitivity and high specificity of targeted DENV NS1 protein, so that the clinical detection requirements of DENV can be met.

The invention relates to the technical field of antibodies, in particular to a 3D protein monoclonal antibody and application thereof. The monoclonal antibody comprises a heavy chain variable region and a light chain variable region, the heavy chain variable region comprises the following three complementary determining regions CDR: CDR1 shown as SEQ ID No.5, CDR2 shown as SEQ ID No.6, and CDR3 shown as SEQ ID No.7; the heavy chain variable region comprises the following three complementary determining regions: CDR1 shown as SEQ ID No.5, CDR2 shown as SEQ ID No.6, and CDR3 shown as SEQ ID No.7; the light chain variable region comprises the following three complementary determining regions CDR: CDR1 as shown in SEQ ID No.8, CDR2 with the sequence of WAS, and CDR3 as shown in SEQ ID No.9. The light chain variable region has the advantages that the light chain variable region can be used for preparing the light chain variable region; the invention provides a specific monoclonal antibody of a foot and mouth disease virus non-structural protein 3D protein. A Western blotting test shows that the specific monoclonal antibody specifically reacts with the non-structural protein 3D; indirect immunofluorescence tests show that the antibody has good reactivity with an FMDV O-type strain.

The present disclosure relates to polypeptides that specifically bind to Dengue virus non-structural protein 1, including antibodies and fragments thereof. The antibody or antigen-binding fragment thereof may specifically bind Dengue virus (DENV) serotype 4 and include: a heavy chain variable region that comprises at least one CDR amino acid sequence selected from the group consisting of: SGYNWH, YIHYSGGTNYNPSLKS, RTGTVPFAY, SYVMH, YLNPYNDDTKYNEKFKG, and GPPYALDY. The present disclosure further relates to methods of producing the polypeptides of the present disclosure, methods of diagnosing DENV, and methods of treating a DENV infection.

This invention provides an optimized method for enhancing self-amplifying RNA expression and an enhanced self-amplifying RNA. Specifically, it provides an optimized method for enhancing self-amplifying RNA expression, which involves codon optimization. The optimized region is a specific region of the alphavirus non-structural protein sequence of the self-amplifying RNA, where the alphavirus non-structural protein is nsP1234. It also provides a method for preparing the enhanced self-amplifying RNA, an enhanced self-amplifying RNA, and a plasmid. This application improves the expression efficiency of the self-amplifying RNA target gene by designing and constructing codon sequence optimization for certain regions of these non-structural proteins.

The present disclosure provides a self-amplifying RNA (sa-mRNA) comprising an open reading frame encoding a replicase or a non-structural protein 4 (nsP4) and an open reading frame encoding peptide or polypeptide, wherein the open reading frame encoding the peptide or polypeptide is co-expressed with the replicase or nsP4.

The invention belongs to the field of gene engineering, and particularly relates to a recombinant bluetongue virus for fusion expression of a tetracysteine (TC) tag on an NS2 protein and a construction method of the recombinant bluetongue virus. The recombinant bluetongue virus BTV1-NS2-354TC is constructed by inserting a TC tag into the C terminal (behind the 354 amino acid) of a wild type bluetongue virus non-structural protein NS2; the biological characteristics of the recombinant bluetongue virus and the biological characteristics of a wild type bluetongue virus are not obviously different, the TC tag-carrying NS2 protein expressed by the recombinant bluetongue virus can be dyed by a double-arsenic dye (FlAsH-EDTA), green fluorescence is displayed under a fluorescence microscope, and the recombinant bluetongue virus can be used for visual quantitative detection of BTV infection and positioning and tracing research of the NS2 protein.

The invention discloses a prawn culture pathogenic bacteria multiple ERA detection primer and probe based on a biochip technology and application of the prawn culture pathogenic bacteria multiple ERA detection primer and probe. The kit is characterized in that the primers and the probe respectively aim at tcdB genes of highly pathogenic vibrios, hypothetical protein genes of white spot syndrome viruses, 16S rRNA genes of enterocytozoon hepatopenaei, non-structural protein genes of infectious subcutaneous and hematopoietic necrosis viruses, DNA primer enzyme genes of decapod iridovirus 1 and PirB genes of bacterial prawn acute hepatopancreatic necrosis pathogens; the method comprises the following steps of: detecting a capsid protein gene of a prawn infectious muscle necrosis virus, a capsid protein gene of a nedavirus, a capsid protein gene of the nedavirus, a structural protein ORF1 gene of the prawn infectious muscle necrosis virus, a non-structural polyprotein gene of a taura syndrome virus and a Rep-1b gene of a yellow head virus, and further discloses specific sequences. The sensitivity, the specificity and the accuracy are high.

The present disclosure provides novel self-amplifying RNA (saRNA) constructs that demonstrate enhanced protein expression, prolonged durability, reduced immunogenicity, and the ability to express multiple therapeutic proteins homogeneously. The saRNA constructs comprise a 5' untranslated region (5'UTR), non-structural protein genes derived from alphaviruses, at least one gene of interest encoding a therapeutic protein, a 3' untranslated region (3'UTR), and one or more modified nucleosides. Also disclosed are dual construct systems comprising a first construct encoding non-structural proteins and a second construct encoding one or more genes of interest. Methods of producing and using the saRNA constructs for engineering cells, particularly immune cells, for treatment of various conditions including cancer, inflammatory conditions, and infectious diseases are provided. The saRNA constructs enable the generation of "armored" immune cells expressing multiple therapeutic proteins, thereby providing a multi-pronged approach to complex diseases.

The present application relates to a method for extracting epitopes effective in the prevention or treatment of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), the method comprising the steps of: (a) extracting, from an anti-spike antibody database, first structural data including known structures and second structural data including unknown structures, and extracting, from a SARS-CoV-2 proteome database, third structural data including known structures; (b) carrying out 3D modeling on the second structural data, and classifying the third structural data into a first group including structural proteins and a second group including both structural proteins and non-structural proteins; (c) carrying out spike-antibody docking on the basis of the first group and the second structural data on which 3D modeling has been carried out; (d) constructing a spike-antibody database on the basis of the first structural data and the docked data, and predicting conformational epitopes on the basis of the second group; (e) characterizing epitopes on the basis of the spike-antibody database and the conformational epitopes; and (f) selecting final epitopes on the basis of analysis results.