35 results about "Site-specific recombination" patented technology

Among other things, provided herein are systems that replace the natural random integration activity of a retrovirus with site-specific integration machinery. This approach allows for a more precise targeting of a gene of interest into a human genome, e.g., for therapeutic purposes. The system may include integration-deficient retrovirus (e.g., lentivirus) (IDLV), in which the natural integration activity has been reduced (e.g., by mutation to the viral integrase polypeptide). Instead, the system may comprise a site-specific recombinase (e.g., a serine recombinase, e.g., a serine integrase) capable of directing insertion of a template DNA, or portion thereof, into a desired site in the human genome.

Disclosed herein are engineered LSR sequences, compositions, and uses including in genome-editing systems and therapeutic compositions. The LSRs can facilitate nucleic acid recombination at particular attachment sites, using suitable donor sequences. The disclosed LSRs may be used to site-specifically recombine or integrate nucleic acid sequences for a variety of purposes, including treatment of human diseases.

A homologous recombination method of excising a specific DNA sequence from a DNA molecule. The method includes providing the DNA molecule including a pair of target sequences that are a target of a site-specific recombination enzyme and a pair of homologous sequences that are DNA sequences sharing homology with each other. The pair of target sequences are oriented in different directions from each other and arranged on opposite sides of the specific DNA sequence. The pair of homologous sequences are arranged on opposite sides of the specific DNA sequence at positions further outside of the pair of target sequences. The method further includes inducing homologous recombination between the pair of homologous sequences by action of the site-specific recombination enzyme on the DNA molecule so that the specific DNA sequence and the pair of target sequences are excised from the DNA molecule.

Disclosed are altered recombinases wherein at least one amino acid is different from a parent, wildtype recombinase and the altered recombinase has improved recombination efficiency towards wild-type and / or pseudo att site sequences relative to the parent, wildtype recombinase. Disclosed are altered modular recombinases comprising binding (or interacting) regions from a different recombinase. Disclosed are methods of modifying the genomes of cells using the altered recombinases, including methods of site-specifically integrating a polynucleotide sequence of interest in a genome of a eucaryotic cell.

The present invention relates generally to the field of site-specific DNA recombination mediated by lambda integrases, and more specifically to methods of producing eukaryotic cells and cell lines comprising a genomic landing pad for lambda integrase mediated recombination, as well as the eukaryotic cells themselves and subsequent methods of their use for lambda integrase mediated recombination and as bioreactors for cell therapies.

In a homologous recombination method in which a specific DNA sequence is deleted from a DNA molecule, the DNA molecule is used as the DNA molecule. A DNA molecule is prepared in which a pair of target sequences, which are targets for a site-specific recombinase, are disposed so as to sandwich a specific DNA sequence therebetween and so as to have different orientations from each other, and a pair of homologous sequences, which are DNA sequences having homology, are disposed on the outside of the pair of target sequences disposed so as to sandwich the specific DNA sequence. The site-specific recombinase is caused to act on a DNA molecule, and a specific DNA sequence and a pair of target sequences are deleted from the DNA molecule by homologous recombination between a pair of homologous sequences.

Recombinant DNA molecules and constructs are provided that are useful for modulating gene expression in plants. One or more expression cassette(s) of a recombinant DNA molecule or construct may be excised from transgenic plants following transformation by the presence of flanking site-specific recombination sites in the recombinant DNA molecule or construct by expression of a recombinase enzyme encoded by the recombinant DNA molecule or construct. Such a recombinase system may be used to remove such expression cassette(s) from plants transformed with the recombinant DNA construct or vector. The recombinase transgene may be operably linked to a tissue-preferred or tissue-specific promoter for autoexcision in transformed plants without crossing to a different transgenic line expressing the recombinase. Methods for causing autoexcision of one or more expression cassette(s) in a transgenic plant, and plants and cells containing or transformed with a recombinant DNA molecule or construct of the present disclosure, are also provided.

The present invention relates to a method for integrating one or more recombinant sites in a sole target gene of a cell genome comprising carrying out sequentially multiple site-specific recombination by using alternatively two different serine-integrases, wherein the selection marker is removed after each site-specific recombination by different transposases: An in vitro method for inserting at least one recombinant site in a target gene / locus from an isolated cell genome, said method comprising integrating a nucleotide sequence A into the target gene from an isolated cell genome by homologous recombination, wherein said nucleotide sequence A comprises a recombination cassette, flanked at 5′ and 3′ by target gene homology arms, and removing the selection marker by using the second transposase, thus obtaining a cell genome A comprising (i), (ii) and (iii) of nucleotide sequence A.

Recombinant DNA molecules and constructs are provided that are useful for modulating gene expression in plants. One or more expression cassette(s) of a recombinant DNA molecule or construct may be excised from transgenic plants following transformation by the presence of flanking site-specific recombination sites in the recombinant DNA molecule or construct by expression of a recombinase enzyme encoded by the recombinant DNA molecule or construct. Such a recombinase system may be used to remove such expression cassette(s) from plants transformed with the recombinant DNA construct or vector. The recombinase transgene may be operably linked to a tissue-preferred or tissue-specific promoter for autoexcision in transformed plants without crossing to a different transgenic line expressing the recombinase. Methods for causing autoexcision of one or more expression cassette(s) in a transgenic plant, and plants and cells containing or transformed with a recombinant DNA molecule or construct of the present disclosure, are also provided.

The invention relates generally to the field of genome editing and provides DNA recombinases, which efficiently and specifically recombine genomic target sequences via the fusion of recombinase monomers. More specifically, the invention provides a method to generate a fusion protein for efficient and specific genome editing, comprising a complex of recombinases comprising at least a first recombinase enzyme, a second recombinase enzyme and at least one linker, wherein said first recombinase enzyme and said second recombinase enzyme specifically recognize a first half-site and a second half-site of an upstream target site and / or a downstream target site of a recombinase; wherein said first recombinase enzyme and said second recombinase enzyme are interconnected via a linker; and wherein said linker comprises or consists of an oligopeptide. The invention further relates to fusion proteins generated with this method. The invention also discloses designer-recombinases, which catalyze the inversion of a DNA sequence present in the int1h regions on the human X chromosome. The invention further relates to nucleic acid molecules encoding said DNA recombinases and fusion proteins, as well as to the use of said fusion proteins, DNA recombinases and nucleic acid molecules in a pharmaceutical composition.

Disclosed herein are nanorod productions systems (NPS) useful for the production of biological scalable functionalization-ready nanorods (BSFnano). The nanorods produced are derived from filamentous phage Ff (f1, M13 or fd). The NPS disclosed herein permits efficient biological production of non-infectious, heat-stable isomorphic proteinaceous nanorods comprising modifications allowing site-specific recombinant, chemical and enzymatic attachment of peptide and non-peptide functionalities in an orthogonal manner. Also disclosed are methods of making and using these nanorods, such as in methods of detecting target molecules.

Recombinant DNA molecules and constructs are provided that are useful for modulating gene expression in plants. One or more expression cassette(s) of a recombinant DNA molecule or construct may be excised from transgenic plants following transformation by the presence of flanking site-specific recombination sites in the recombinant DNA molecule or construct by expression of a site-specific recombinase enzyme encoded by the recombinant DNA molecule or construct. Such a recombinase system may be used to remove expression cassette(s) from plants transformed with the recombinant DNA construct or vector. The recombinase transgene may be operably linked to a promoter suitable for autoexcision in transformed plants without crossing to a different transgenic line expressing the recombinase. Methods for causing autoexcision of one or more expression cassette(s) in a transgenic plant, and plants and cells containing or transformed with a recombinant DNA molecule or construct of the present disclosure, are also provided.

Disclosed are altered recombinases wherein at least one amino acid is different from a parent, wildtype recombinase and the altered recombinase has improved recombination efficiency towards wild-type and / or pseudo att site sequences relative to the parent, wildtype recombinase. Disclosed are altered modular recombinases comprising binding (or interacting) regions from a different recombinase. Disclosed are methods of modifying the genomes of cells using the altered recombinases, including methods of site-specifically integrating a polynucleotide sequence of interest in a genome of a eucaryotic cell.

The application discloses a corynebacterium striatum phage and application thereof. The corynebacterium striatum phage is a corynebacterium striatum phage CSP1, the preservation number of which is CGMCC NO: 45598, and the preservation time is July 4, 2023. The corynebacterium striatum phage has strong tolerance, biological safety and carries site-specific recombination functional elements (integrase and recombination sites). Meanwhile, the corynebacterium striatum phage can lyse 20 strains of multi-drug resistant corynebacterium striatum, and lays a foundation for subsequent medical researches using the corynebacterium striatum phage as a test and treatment carrier, a targeted pathogenic bacterium or symbiotic bacterium, and an immune regulator inducer.

The invention discloses a recombinase-based highlight induced photosynthetic electron transfer loop and application thereof, the loop uses a highlight induced ELIP1 promoter to drive FLP recombinase expression, and regulates and controls the expression of a photosynthetic electron transfer key component gene PETE or PETC through an FLP / FRT site specific recombination system. The problem of light inhibition caused by overload of a plant photosynthetic electron transfer chain in a highlight environment is solved, light intensity dependent gene expression regulation and control are realized, the photosynthetic electron transfer characteristic under highlight stress is effectively improved, and the photoadaptability and photosynthetic efficiency of plants are enhanced. The system provides a new tool for crop stress resistance molecular breeding and synthetic biology research.

The present disclosure relates to a novel screening method for screening a plurality of pGREs in parallel and nucleotide constructs for use in the method. The construction body comprises a nucleotide block, the sequence of the nucleotide block is PRIMREG1-pGREE-PRIMREG2-GENEE or GENE-PRIMREG1-pGREE-PRIMREG2, and the PRIMREG1 is a primer recognition sequence 1 or an FLEx box containing the primer recognition sequence 1. PGRE is a presumed gene regulatory element, PRIMREG2 is a primer recognition sequence 2 or a FLEx cassette comprising a primer recognition sequence 2, GENE comprises nucleotide blocks REPSEQ, IRES and SSR, where REPSEQ is a reporter gene and is present or absent, IRES is an internal ribosome entry site or a 2A self-cleaving peptide, and SSR is a site-specific recombinase gene. At least one of the PRIMREG1 and the PRIMREG2 is a FLEx cassette comprising a primer recognition sequence.

Recombinant DNA molecules and constructs are provided that are useful for modulating gene expression in plants. One or more expression cassette(s) of a recombinant DNA molecule or construct may be excised from transgenic plants following transformation by the presence of flanking site-specific recombination sites in the recombinant DNA molecule or construct by expression of a site-specific recombinase enzyme encoded by the recombinant DNA molecule or construct. Such a recombinase system may be used to remove expression cassette(s) from plants transformed with the recombinant DNA construct or vector. The recombinase transgene may be operably linked to a promoter suitable for autoexcision in transformed plants without crossing to a different transgenic line expressing the recombinase. Methods for causing autoexcision of one or more expression cassette(s) in a transgenic plant, and plants and cells containing or transformed with a recombinant DNA molecule or construct of the present disclosure, are also provided.

The invention belongs to the technical field of bioengineering, and particularly relates to a recombinant plasmid for promoting synthesis of tylosin, an engineering bacterium and a preparation method thereof, the genetically engineered bacterium (Streptomyces fradiae G3-F2) is preserved in China Center for Type Culture Collection on February 24, 2025, and the preservation number is CCTCC NO: M 2025278. According to the streptomyces fradiae G3-F2 disclosed by the invention, through site-specific recombination, a recombinant plasmid is constructed by utilizing an optimized ZouA gene DNA amplification system, so that multi-copy increase of a tylF gene is realized, and further, streptomyces fradiae G3-F2 is prepared; the enzymatic reaction rate of the methyltransferase is greatly improved; the efficient conversion of the component C is realized; and low-temperature culture fermentation is combined, so that the tylosin synthesis rate and efficiency are greatly improved. The titer of the strain in a 50L tank can reach 18500U / mL under the low-temperature culture condition of 28 DEG C, so that the production cost can be greatly reduced.

The application discloses a method for continuous editing of multi-site chromosomes in eukaryotic cells. The application provides a method for integrating the genome of eukaryotic cells, comprising: using a knock-in site-specific recombination site (such as noxM) in the genome of eukaryotic cells, realizing integration of a large fragment circular DNA molecule at a specific site of the genome under the action of a knock-in site-specific recombinase (such as Nigri), and simultaneously using an adjacent knock-out site-specific recombination site (such as loxP or a mutant thereof), rearranging the related sites under the action of a knock-out site-specific recombinase (such as Cre), so as to realize continuity of integration. The novel editing platform of high-efficiency site-specific recombination provided by the application can realize directional, continuous and multi-site integration of a large fragment DNA in eukaryotic cells, and the application has important application prospects in the field of biological technologies including plant breeding, animal cell line development, fungus engineering and the like.

This invention belongs to the field of bioengineering technology, specifically relating to a recombinant plasmid for tylosin synthesis, an engineered bacterial strain, and its preparation method. The genetically engineered bacterial strain (Streptomyces freundii G3-F2) was deposited at the China Center for Type Culture Collection (CCTCC) on February 24, 2025, with accession number CCTCC NO: M 2025278. This invention achieves site-specific recombination, utilizes an optimized ZouA gene DNA amplification system to construct the recombinant plasmid, and realizes… tulle Multiple copies of the F gene were increased, resulting in the preparation of *Streptomyces freundii* G3-F2. This significantly improved the rate of methyltransferase enzymatic reaction and achieved efficient conversion of component C. Combined with low-temperature fermentation, the rate and efficiency of tylosin synthesis were greatly enhanced. The strain achieved a titer of 18500 U / mL in a 50L tank under 28℃ low-temperature culture conditions, which can significantly reduce production costs.

The application provides a tumor cell dormancy and awakening marker tracing system (DormTracer), specifically, the system combines a gene switch and a site-specific recombination technology, expression of a reporter gene is driven by a recombination enzyme fused with a dormancy marker protein, the reporter gene is affected by a cell dormancy history, but does not affect the cycle of cell dormancy and proliferation, therefore, the tumor cell dormancy history can be traced, and the current instantaneous state of the cell is not limited to monitoring. The system can distinguish tumor cells in three states of proliferation, dormancy and dormancy awakening.

The invention relates to a method for tracing mononuclear macrophages. Comprising the following steps: placing a coding sequence of a fusion protein formed by connecting a site-specific recombinase and a ligand binding domain which can be combined with an exogenous ligand and regulate and control the activity of the recombinase after combination under the control of a regulation and control element of a C3aR1 gene, so that the expression of the fusion protein is limited in a cell with a C3aR1 expression history; setting an excidable blocking sequence corresponding to the recombinase recognition site in a genome of the recombinase recognition site, so that the blocking sequence inhibits the expression of a signal output molecule when the recombinase recognition site is not recombined; after the exogenous ligand is given, the recombinase is activated by regulating and controlling the conformational change of the ligand binding domain in the fusion protein, so that the recombinase is cut off the resectable blocking sequence, and the inhibition on the signal output molecule is relieved; and the signal output molecule for tracing is irreversibly expressed in the mononuclear macrophage with the C3aR1 expression history.

A fast and accurate three-plasmid oncolytic adenovirus recombinant packaging system Ad5MixPlus and an application thereof are provided. The system is composed of three adenovirus recombinant plasmids. The core technology of the system is that two sets of different site recombination sequences are skillfully loaded on a first 5-type adenovirus right arm backbone plasmid large vector, then two small shuttle plasmids respectively provide a right arm-modified Hexon / E3 / Fiber sequence and an E1a expression cassette controlled by a left arm tumor-specific promoter, and the difficulties and obstacles to the modification of the adenovirus backbone large vector are overcome. After two rounds of site-specific recombination, the ideal oncolytic adenovirus is packaged accurately and quickly.

The present disclosure relates to a method for preparing a site-specific recombination polynucleotide molecule derived from the attP site of the bacteriophage mv4 and to a kit for such site-specific recombination. The kit can be used to transform procaryote hosts to integrate any polynucleotide sequence of interest.

Compositions and methods are provided for the introduction and the regulated expression of genes in plants. Compositions include promoter constructs that provide a level of activity useful for the regulated expression of site-specific recombinases, while avoiding premature excision. Further provided are isolated polynucleotides encoding novel babyboom polypeptides, expression cassettes, and plants comprising the same. Methods for the introduction of genes into plants are provided, including methods for plastid transformation and methods for the transformation of tissues from mature seeds and leaves.

Compositions and methods are provided for the introduction and the regulated expression of genes in plants. Compositions include promoter constructs that provide a level of activity useful for the regulated expression of site-specific recombinases, while avoiding premature excision. Further provided are isolated polynucleotides encoding novel babyboom polypeptides, expression cassettes, and plants comprising the same. Methods for the introduction of genes into plants are provided, including methods for plastid transformation and methods for the transformation of tissues from mature seeds and leaves.

Disclosed herein are engineered LSR sequences, compositions, and uses including in genome-editing systems and therapeutic compositions. The LSRs can facilitate nucleic acid recombination at particular attachment sites, using suitable donor sequences. The disclosed LSRs may be used to site-specifically recombine or integrate nucleic acid sequences for a variety of purposes, including treatment of human diseases.

The present invention relates to site-specific DNA recombinase enzymes comprising a nuclear localisation signal (NLS). More specifically, the present invention relates to a Bxb1 serine integrase comprising an NLS and methods of use thereof, in particular for antibody display and expression

The present invention is related to a nucleic acid molecule, which is also referred to as third nucleic acid molecule, wherein the third nucleic acid molecule comprises(1) a nucleic acid molecule comprising the following elements:(a) optionally, a first part of a genome of a virus;(b) a nucleotide sequence, preferably a genomic nucleotide sequence, or a transcription unit;(c) a regulatory nucleic acid sequence which has a regulatory activity in a prokaryote;(d) exactly one site-specific recombination site;(e) a nucleotide sequence providing for a negative selection marker;(f) a bacterial nucleotide sequence unit comprising (i) bacterial nucleotide sequences for conditional replication and (ii) a nucleotide sequence providing for a positive selection marker;(g) optionally a first restriction site; or(2) a nucleic acid molecule comprising a nucleotide sequence according to SEQ ID NO: 6; or(3) a nucleic acid molecule identical or similar to the nucleic acid molecule contained in the organism deposited with the DSMZ under the Budapest treaty under accession number DSM 23754, wherein preferably the nucleic acid molecule contained in the organism is a heterologous nucleic acid molecule;wherein the third nucleic acid molecule is either a linear or a circular molecule.

The invention relates to methods and systems for gene editing combining endonuclease mediated homology directed repair using single stranded oligonucleotides templates with a site-specific recombinase to allow large cargos to be integrated at any location in the genome.