571 results about "Expression cassette" patented technology

The invention discloses an aspergillus niger recombinant bacterium AnCT-XynB-K-agdA and application thereof, and belongs to the technical field of microbiology and bioengineering. According to the invention, Aspergillus niger AnCat is taken as an expression host, firstly, an acid protease regulatory factor prtT is knocked out to obtain a defective strain, and then different chaperonins are subjected to fusion expression by optimizing integration sites of a laccase lcc9 expression cassette, so that the high-expression laccase chaperonin is obtained. According to the method, an expression vector pC3-5 'agdA-PcitA-XynB-L-lcc9-hph-3' agdA is constructed, a recombinant bacterium AnCT-XynB-K-agdA is further obtained, the enzyme activity of the extracellular laccase subjected to shake flask fermentation reaches 1821.2 U / L, and finally, the recombinant efficient expression of the laccase Lcc9 in aspergillus niger is realized.

The invention provides a structure, a sequence and a construction method of a plant virus gene editing vector and application of the plant virus gene editing vector in plant gene editing. The method is characterized in that a TRV2 genome of a tobacco embrittlement virus (TRV) is used for carrying and expressing a small Cas gene AsCas12f and gRNA of the small Cas gene AsCas12f. Preferably, the AsCas12f and the gRNA of the AsCas12f are subjected to tandem expression by utilizing the TRV2 to form an expression cassette structure of the AsCas12f-tRNA-gRNA, the expression cassette substitutes a part of 2b gene sequence of the TRV2 on a TRV2 genome to form a plant gene editing vector pTRV2-Cas12f-gRNA based on the TRV virus, and long-time stable expression and gene editing can be performed after wild type nicotiana benthamiana is inoculated. The invention also provides a method for carrying out gene editing in a wild type plant by utilizing the plant virus.

The invention provides a system and a method for screening Cas protein mutants capable of identifying different PAM sequences, and belongs to the technical field of bioengineering. The system comprises a Cas protein mutant screening vector and an sgRNA expression vector, the Cas protein mutant screening vector comprises the following expression elements: a Cas protein coding gene, a plasmid replicon and a first resistance screening tag expression cassette; the sgRNA expression vector comprises the following expression elements: a plasmid replicon, a second resistance screening tag expression cassette, a third resistance screening tag expression cassette and an sgRNA expression cassette. According to the system and the method, aiming at the problem that the efficiency of identifying PAM sequences except NGG by wild type spCas9 protein is low, an active Cas9 mutant capable of identifying PAM sites except NGG is screened out, and the application field and the editing efficiency of a CRISPR gene editing system can be greatly expanded.

The present invention provides a novel therapeutic drug for treating a Prom1-associated retinal disease. Specifically, the present invention provides an optimized Prom1 gene expression cassette, an rAAV viral vector, and a gene therapy drug. The drug of the present invention can specifically express the PROM1 protein in the retinal photoreceptor layer, and is suitable for the clinical treatment of a retinal disease associated with Prom1 gene mutation.

This invention provides vectors (e.g., lentiviral vectors) and expression cassettes for incorporation into vectors for expressing an alpha-globin (α-globin) gene. In certain embodiments method of treatment using such vectors are provided.

96 New PCT-Patent Application based on EP 24 186 635.9 Insempra GmbH Vossius Ref.: AG4141 PCT S3 Abstract The present invention relates to a linear double-stranded deoxyribonucleic acid (dsDNA) molecule comprising one or more Tus protein (Tus) binding site(s) at the 5'-terminus, one or more Lac repressor protein (LacI) binding site(s) at the 3'-terminus, and a segment comprising a DNA sequence of interest (DOI) between said Tus binding site(s) and said LacI binding site(s). The present invention further relates to a non-naturally or naturally occurring RNA molecule as encoded by or transcribable / transcribed from the linear dsDNA molecule of the invention, and to an expression DNA cassette comprising a promoter, an RBS, a GOI encoding a POI, and one or more LacI binding site(s), wherein said expression cassette does not comprise a terminator sequence between the 3´ end of said GOI and said one or more LacI binding site(s). The present invention also relates to a non-naturally or naturally occurring RNA molecule as encoded by or transcribable / transcribed from said expression cassette. The present invention further relates to method of protecting a linear deoxyribonucleic acid (DNA) molecule from exonuclease degradation by adding one or more Tus binding site(s) at the first terminus of the DNA molecule and adding one or more LacI binding site(s) at the other terminus of the DNA molecule. The present invention further relates to a method of synthesizing a protein of interest (POI) in a cell- free protein synthesis (CFPS) reaction mixture by using the (ds)DNA and / or RNA molecules of the invention, and also to a cell-free biological system or CFPS reaction mixture comprising the (ds)DNA, expression cassette and / or RNA molecules of the invention.

The invention discloses a method for improving the solubility and thermal stability of sweet protein, and belongs to the technical field of biosynthesis, the method comprises the following steps: constructing a gene tandem recombinant plasmid containing a monellin x-3C-sfGFP-3C-monellin y expression cassette, x is greater than or equal to 1, y is greater than or equal to 1, 5 is greater than or equal to x + y is greater than or equal to 3, a 3C protease recognition sequence is also inserted between adjacent copies of monellin, and each monellin is connected with a purification tag; the gene tandem recombinant plasmid is transformed into escherichia coli and inducible expression is carried out, thalli are split and purified to obtain fusion protein, 3C protease is used for enzyme digestion, and the target protein is obtained after re-purification. The total protein yield and the solubility proportion are remarkably improved by adopting a series construction mode of plasmids, meanwhile, the stability of monellin is effectively improved, the purification step is simpler and more convenient, and the method is suitable for industrial production.

Herbicide-resistant rice plants, isolated polynucleotides that encode herbicide resistant and wild-type acetohydroxy-acid synthase large subunit 1 (AHASL1) polypeptides, and the amino acid sequences of these polypeptides, are described. Expression cassettes and transformation vectors comprising the polynucleotides of the invention, as well as plants and host cells transformed with the polynucleotides, are described. Methods of using the polynucleotides to enhance the resistance of plants to imidazolinone herbicides, and methods for controlling weeds in the vicinity of herbicide-resistant plants are also described.

The present disclosure relates to a CCL20 promoter derived from a regulatory region of the CCL20 gene, which stimulates gene expression in response to inflammation. These CCL20 promoters, which can be stimulated by endogenous or exogenous cytokines, can be used to control the timing of expression of heterologous genes. The disclosure also relates to expression cassettes or vectors comprising a CCL20 promoter of the disclosure operably linked to a nucleic acid sequence encoding, for example, a polypeptide of interest; and delivery systems (e.g., viral particles, lipid vesicles or nanoparticles) or cells comprising such expression cassettes or vectors. The disclosure also relates to the use of the promoter, expression cassette, vector, delivery system or cell in the treatment of inflammation-related diseases or for recombinant gene expression.

Provided herein are modified nucleic acids comprising modified MTM1 genes, modified muscle-specific promoters, e.g., MHCK7, or a combination thereof. Also provided herein are expression cassettes, AAV vector genomes, and AAV particles comprising the same. Further provided herein are methods of delivering MTM1 to a muscle cell and methods of treating XLMTM.

The invention provides a novel therapeutic drug for treating Prom1 related retinal diseases. Specifically, the invention provides an optimized Prom1 gene expression cassette, an rAAV virus vector and a gene therapy drug. The medicine provided by the invention can specifically express the PROM1 protein in a retina photoreceptor layer, and is suitable for clinical application to treatment of Prom1 gene mutation related retinal diseases.

The invention provides a chicken PGC reporter gene cell strain for detecting the biological activity of FGF2 as well as a construction method and application of the chicken PGC reporter gene cell strain, and belongs to the technical field of biological activity detection. The cell strain is a recombinant cell which is stably integrated with a reporter gene expression cassette responding to an FGF signal channel in a chicken PGC genome; the expression cassette comprises a serum response element (SRE), a minimum promoter and a reporter gene (such as a luciferase gene). The construction method comprises the following steps: introducing a donor vector containing the elements into chicken PGC by utilizing a PiggyBac transposon system, and screening to obtain a stably expressed cell strain. The cell strain provided by the invention retains endogenous signal channel characteristics of chicken PGC, and can specifically and highly sensitively respond to FGF2 stimulation. The method can be used for biological activity quantitative determination and thermal stability evaluation of the FGF2 protein and the mutant thereof, quality control of culture medium components and screening of an FGF signal channel regulator.

The present invention relates to a recombinant construct comprising: a first expression cassette comprising a fluorescence enhancing factor; wherein the amino acid sequence of the fluorescence enhancement factor is as shown in SEQ ID NO: 2, or an amino acid sequence which at least has 60%, 65%, 70%, 75%, 80%, 85% or 90% identity with the SEQ ID NO: 2 and is functionally equivalent to the protein as shown in the SEQ ID NO: 2; the first expression cassette is configured to increase the self-luminous intensity of a plant, plant tissue, or cell. According to the application, the LumEnhancer gene is further introduced on the basis of an FBP biological self-luminous system, so that the physiological activity of transgenic plant cells is effectively improved, and the energy metabolism level of the plant is further enhanced, thereby providing a more sufficient energy basis for a bioluminescence reaction and effectively enhancing the luminous intensity of the whole plant.

The present invention relates to compositions comprising an RNA molecule and pharmaceutical formulations comprising such compositions wherein the RNA molecule comprises an expression cassette encoding an immunogenic peptide comprising at least two fragments wherein each fragment comprises at least one epitope wherein the epitope is derived from an amino acid sequence encoded by human immunodeficiency virus (HIV). In addition, the present invention relates to a method of preventing or treating HIV.

Disclosed herein is a promoter sequence comprising at least a portion of a ABCB4 promoter sequence or reverse complement thereof, wherein the splice sites of the ABCB4 promoter are mutated, and lentiviral vector sequences comprising said promoter sequences operably linked to a transgene. Preferably, the promoter sequence and transgene are present in a reverse expression cassette in reverse orientation to the direction to the lentiviral vector sequence. When a transgene for ABCB4 is used, the lentiviral vector can be used in the treatment of PFIC3.

The present application relates to the field of genetic engineering, and in particular to a vector for analyzing plant promoter expression specificity, a preparation method and application thereof. The T-DNA region of the vector comprises, from 5' end to 3' end, a callus-specific promoter-driven selection marker gene expression cassette and a reporter gene expression cassette with a multiple cloning site; the callus-specific promoter-driven selection marker gene expression cassette comprises a callus-specific promoter, a selection marker gene coding sequence and a terminator. The vector of the present application utilizes a rice callus-specific promoter to drive the expression of a selection marker gene, can greatly reduce the non-specific interaction with the promoter of a target gene, improve the specificity of the expression of the target gene, and effectively reduce the biological safety risk caused by the selection marker gene in a transgenic plant, and has important application value in basic theoretical research and molecular breeding.

This invention relates to the fields of bioengineering and fermentation engineering, and discloses a genetically engineered *Streptomyces albopictus* strain that produces high levels of ε-polylysine and its fermentation production process. This genetically engineered strain uses *Streptomyces albopictus* strains acclimated to ε-polylysine tolerance as a chassis, and its genome integrates... ppc and dapf Through expression box, heterogeneous asd Expression cassettes, containing secretory signal peptides lysp Fusion expression cassettes and D404A / K499A point mutations pls Expression cassette. The fermentation process employs a two-stage pH control strategy, maintaining a neutral environment during the cell growth phase and adjusting to an acidic environment during product synthesis. This invention improves ε-polylysine yield through a synergistic strategy of enhancing precursor supply, constructing efflux detoxification channels, and increasing synthase activity; combined with an acidic fermentation process, it effectively inhibits product degradation, yielding a high-molecular-weight and highly uniform target product suitable for industrial production.

The invention provides a recombinant herpes simplex virus and a herpes simplex virus modification method. The herpes simplex virus modification method comprises the step of replacing an ICP4 protein coding gene in a herpes simplex virus genome containing an infected cell protein 4 (ICP4) gene with an hTERTp-fluorescent protein expression cassette. The expression cassette comprises an hTERTp promoter and a fluorescent protein coding sequence controlled by the hTERTp promoter, and the transcription direction is opposite to that of an ICP4 promoter in a genome. The hTERTp-fluorescent protein expression cassette in the recombinant virus obtained by the method disclosed by the invention can be normally expressed in response to telomerase activity. Therefore, the recombinant virus is capable of expressing a fluorescent protein, such as mBaoJin, in a cell having human telomerase activity. Cells infected by the virus can be identified through fluorescence signals, and the higher the telomerase activity is, the stronger the fluorescence intensity is. The virus has wide application value in research of tumor action mechanisms and stem cell action mechanisms, health assessment, screening of tumor drugs, research and development of diagnostic reagents and establishment of animal models.

This invention relates to polynucleotides comprising UBE3A open reading frame (ORF) sequences, vectors comprising the same, and methods of using the same for delivery of the ORF to a cell or a subject and to treat disorders associated with aberrant expression of a UBE3A gene or aberrant activity of a UBE3A gene product in the subject, such as Angelman Syndrome.

This application relates to nucleic acid regulatory elements (NAREs) that can enhance gene expression in the central nervous system. This application further relates to methods utilizing NAREs and uses of NAREs. Expression cassettes and vectors containing NAREs are also disclosed. These are particularly useful for applications using gene therapy.

The invention relates to a method for splitting, assembling and site-specific integration of large-fragment exogenous DNA (deoxyribonucleic acid) in Trichoderma reesei. Comprising the following steps: (1) constructing an arg:: Ptcu-Cas9-TtrpC-hph strain containing a Cas9 protein; (2) constructing a pUC19 <-> integration site: a pyr4 plasmid; (3) obtaining a plurality of target gene segments containing integration site homologous arms and having overlapping regions at the tail ends of adjacent DNA segments; (4) obtaining a gRNA expression cassette of a targeted genome integration site; (5) arg conversion: the arg is converted into a Ptcu-Cas9-TtrpC-hph strain; and (6) screening and verifying. According to the method, the CRISPR / Cas9 system is combined with the TAR in the trichoderma reesei for the first time, the exogenous large-fragment DNA is split into a plurality of small fragments, then the plurality of small-fragment exogenous DNA are assembled in the trichoderma reesei and are subjected to fixed-point integration, one-step editing of the large-fragment exogenous DNA is realized, and compared with a traditional genetics method, the gene editing efficiency in the trichoderma reesei is greatly improved.

The present disclosure provides a polynucleotide comprising an expression cassette of a membrane protein linked to one or more additional expression cassettes for expressing one or more additional genes, the expression cassette of the membrane protein comprising a transmembrane domain and an extracellular domain, the extracellular domain comprising a recognition sequence, and one or more additional expression cassettes for expressing one or more additional genes, the one or more additional expression cassettes being linked to the one or more additional expression cassettes. The recognition sequence can be specifically recognized by the antibody of the recognition sequence. The invention also provides a vector and a cell containing the polynucleotide, and a method for transferring an exogenous gene into a cell and screening the cell by using the polynucleotide or the vector.

The present invention pertains to the use of miRNA technology for improving recombinant production of polypeptides of interest in host cells. Expression cassettes are provided which produce a miRNA targeting and down-regulating a host cell protein which interferes with production of the polypeptide of interest.

Provided are an anti-DLL3 antibody or an antigen-binding fragment thereof, a chimeric antigen receptor and a mutant thereof, and an enhanced chimeric antigen receptor comprising a PDL1 antagonist and an mIL7 element. Further provided are a nucleic acid encoding same, an expression cassette, vector and cell comprising the nucleic acid, a preparation method, and a use for preventing, treating, detecting, or diagnosing diseases related to DLL3. In the enhanced DLL3 chimeric antigen receptor, the PDL1 antagonist and the cell membrane IL7 cytokine element play a critical role in a long-term anti-tumor process. Animal efficacy experiments have shown complete tumor regression in all mice, indicating broad application prospects in the pharmaceutical field.

Variants of coagulation factor VIII (FVIII) and expression cassettes encoding the FVIII variants thereof are described. A variant FVIII includes a glycoepitope of the FVIII protein including an N2118Q mutation. The N2118Q mutation can be combined with other mutations including a BDD-FVIII, N6, V3, RH, furin-cleavage site deletion. X10, K12, and / or F309S mutation to form additional FVIII variants. The FVIII variants with the N2118Q mutation and expression cassettes thereof can result in reduced immunogenicity of the resulting protein. When combined with other FVIII mutations, higher gene expression, increased secretion, increased stability, and higher FVIII functional activity can be achieved by the expressed FVIII variants. The variant FVIII and expression cassettes described here can be useful in protein replacement therapy and / or gene therapy for the treatment of hemophilia A.

Herein is reported a method for producing a recombinant adeno-associated viral particle preparation (rAAVp), the method comprising the steps of culturing a mammalian cell comprising an expression cassette directed against: a non-adeno-associated viral gene interposed between two AAV inverted terminal repeats (ITRs), and thereby producing the rAAVp; an adeno-associated virus rep gene; an adeno-associated virus cap gene; an adeno-associated virus E1A gene; an adeno-associated virus E1B gene; an adeno-associated virus E2A gene; an adeno-associated virus E4orf6; and an adeno-associated virus VA RNA gene, wherein the culturing is carried out at a pH value in the range of pH 7.4 to pH 7.6 and including a terminal value. The yield of the rAAVp produced by the culture performed at a pH value in the range of pH 7.4 to pH 7.6 and including a terminal value is higher than the yield of the rAAVp produced by the culture performed at a pH value in the range of pH 7.0 to pH 7.2 and including a terminal value, and the yield of the rAAVp produced by the culture performed at a pH value in the range of pH 7.4 to pH 7.6 and including a terminal value is lower than the yield of the rAAVp produced by the culture performed at a pH value in the range of pH 7.0 to pH 7.2. And the rAAVp produced by the culture at a pH value in the range of pH 7.4 to pH 7.6 and including an endvalue has a higher percentage of intact particles than the rAAVp produced by the culture at a pH value in the range of pH 7.0 to pH 7.2 and including an endvalue.

The application discloses an anti-human TRPS1 monoclonal antibody and a preparation method and application thereof, and belongs to the technical field of immunology. The anti-human TRPS1 monoclonal antibody provided by the application comprises VHCDR1, VHCDR2 and VHCDR3 with the amino acid sequences as shown in SEQ ID NO. 1-3, and VLCDR1, VLCDR2 and VLCDR3 with the amino acid sequences as shown in SEQ ID NO. 4-6. The nucleotide sequences of the heavy chain variable region and the light chain variable region of the monoclonal antibody, an expression cassette containing the nucleotide sequences, an expression vector, a recombinant cell and a recombinant bacterium are also provided. Therefore, the monoclonal antibody can be prepared by using genetic engineering technology, and the risk factors in the production and storage process of the traditional monoclonal antibody can be avoided. The monoclonal antibody prepared by the application has stable properties, high experimental repeatability and high titer, and has good specificity and affinity to the TRPS1 protein.

This invention discloses a recombinant vector, recombinant bacteria, and fermentation method for high-yield GHK production, belonging to the field of biomedical technology. The recombinant expression vector of this application comprises a bicistronic expression cassette controlled by a T7 / lac heterozygous promoter, sequentially containing the encoding (GHK). n The vector encodes the first cistron of the polypeptide and the second cistron, where n is an integer from 2 to 200. When this vector is transformed into *E. coli* BL21, the resulting recombinant bacteria can simultaneously express (GHK) after IPTG induction. n Polypeptides and trypsinogen. During fermentation, trypsinogen is activated and enables the processing of (GHK). n In situ enzymatic cleavage of peptides efficiently releases GHK. This invention integrates the traditional multi-step process into a "one-step fermentation," simplifying the production process and reducing purification difficulty. Examples show that the GHK yield can reach up to 3.9 g / L, providing an efficient and convenient new strategy for the large-scale biomanufacturing of GHK.

This invention provides a method for creating a transgenic papaya strain highly resistant to papaya ringspot virus and papaya mosaic virus. The method involves using PRSV and PLDMV of papaya... CP The fusion gene fragment expression cassette and the selection marker gene expression cassette were placed in two separate T-DNA domains and inserted into a plant expression vector. This vector was then introduced into papaya to obtain a papaya line without the selection marker; alternatively, PRSV and PLDMV were used. CP A fusion gene expression cassette and its reverse sequence were tandemly inserted into a plant expression vector, which was then introduced into papaya to obtain... CP A papaya strain with inversely linked gene expression cassettes. The marker-free papaya of this invention does not contain selection markers, effectively improving safety; CP Papaya with inversely linked gene expression cassettes does not require further homozygosity to enhance disease resistance.

Provided is an invention based, in part, on novel gene constructs that encode a microdystrophin protein for use in gene therapy. The microdystrophin gene constructs and expression cassettes were engineered for improved therapy with respect to efficacy, potency and safety to the subject when expressed by a viral vector in muscle cells and / or CNS cells.