39 results about "Genetic transfection" patented technology

The application belongs to the technical field of immunology and in vitro diagnosis, and particularly relates to a rabbit-derived recombinant monoclonal antibody specifically recognizing type II grass carp reovirus VP4 protein, a eukaryotic expression method and application. The S6 gene in GCRV-II encodes VP4 protein, the application transfects the target gene S6 into HEK293 cells for expression and purification, cooperates with Freund's adjuvant to immunize New Zealand white rabbits, and uses ELISA, single B cell screening and eukaryotic expression technology to obtain a rabbit-derived recombinant monoclonal antibody specifically recognizing GCRV-II VP4 protein. The antibody has strong specificity, provides support for further establishment of specific type II grass carp reovirus diagnosis technology, and has great application value for development of GCRV-II related scientific research.

Therefore, this disclosure provides a genetically engineered mesenchymal stem cell (MSC) population, including an expression vector containing the Akt or HGF gene and the PD-L1 gene. It also provides a method for synergistically improving the survival status and immunomodulatory capacity of mesenchymal stem cells or enhancing their proliferation, including transfecting mesenchymal stem cells with the Akt or HGF gene and the PD-L1 gene; and a method for preventing, improving, and / or treating ischemic conditions, enhancing neurogenesis, or reducing neuronal death, including administering an effective amount of the genetically engineered mesenchymal stem cell population of this disclosure to individuals in need.

The application relates to the technical field of biological macromolecular materials, in particular to a preparation method and application of a cationic polymer gene carrier with a fluorinated end group. The cationic polymer gene carrier with a fluorinated end group has the following structure. The fluorinated initiator is used to introduce fluorinated groups in one step to initiate the synthesis of linear polyethylene imine F-PEI with various molecular weights, the introduction of the fluorinated groups improves the gene transfection efficiency, and high-efficiency DNA transfection is realized. The gene carrier can achieve high-efficiency transfection effect in the cell transfection process, the transfection process has small toxicity to cells, gene molecules can be effectively and safely delivered into cells, and the gene carrier has the advantages of high efficiency, low toxicity, low price, simple synthesis and the like. The application has the advantages of simple reaction, easily-obtained raw materials, low preparation cost, easy control of molecular weight, high yield, simple post-treatment and suitability for large-scale production.

The invention provides an electrode suitable for electroporation of early development chick embryos and a preparation method of the electrode. The electrode comprises an upper electrode and a lower electrode; the lower electrode comprises a root end, a middle end and a tail end which are connected with one another; the root end is a straight section which forms an angle of 35-45 degrees with the embryo; the middle end is arc-shaped, and the middle end is in a state of bending downwards and bending forwards at the same time relative to the root end; the tail end is a straight section horizontal to the embryo; the upper electrode is an electrode A suitable for small-range transfection or an electrode B suitable for large-range transfection; the electrode A comprises a straight section and a downward inclined section which are connected with each other and form an angle of 35-45 degrees with an embryo; and the electrode B comprises a straight section, a downward inclined section and a tail section which are connected with one another, wherein the straight section and the embryo form an angle of 35-45 degrees, and the tail section is horizontal to the embryo. By designing a reasonable electrode shape, accurate control of transfection area positioning is facilitated, and gene transfection in an ultra-small target range is realized.

The application relates to the technical field of new biological medical materials, in particular to a cinnamaldehyde modified high-molecular nucleic acid carrier and a preparation method and application thereof, wherein the cinnamaldehyde modified high-molecular nucleic acid carrier is a cinnamaldehyde modified polyethylene imine polymer. The high-molecular nucleic acid carrier provided by the application can significantly improve the DNA and RNA transfection efficiency in different cells, and can effectively improve the biocompatibility of the high-molecular carrier. The preparation steps of the high-molecular nucleic acid carrier are simple, the biocompatibility is good, the repeatability of batches is strong, and the high-molecular nucleic acid carrier has a wide application prospect in the development of nucleic acid carriers, gene transfection and nucleic acid treatment and the like.

The invention belongs to the technical field of biological materials, and particularly relates to an implant coating as well as a preparation method and application thereof. The invention aims to provide a material for an implant coating, which is characterized in that a cytoplasmic membrane and an oxidation response lipid peptide hybrid nanoparticle compress a gene through positive charge adsorption, and the oxidation response lipid peptide hybrid nanoparticle is a lipid peptide formed by connecting a branched basic amino acid head and a hydrophobic tail through an oxidation response bond, the hydrophobic tail part is a long-chain alkyl group or a saturated / unsaturated fatty acid chain, and the cytoplasmic membrane is an amphiphilic cytoplasmic membrane which has high functional protein expression capability after being subjected to hypoxia treatment. The invention provides an innovative combined strategy, namely fusion of gene engineering and bionic technology, and aims to improve the efficiency of coating gene transfection, actively regulate and control the peripheral microenvironment of the implant, accurately promote directional recruitment and enrichment of stem cells and effectively inhibit local inflammatory response, thereby establishing long-term stable bone-implant integration.

The application discloses an electroporation transfection protection fluid suitable for abalone embryo cell gene editing and application thereof, and belongs to the technical field of gene editing. The electroporation transfection protection fluid comprises a basic fluid and a cell protection agent; the basic fluid comprises NaCl, MgSO4, CaCl2, KCl, NaHCO3, NaBr, K2HPO4, glucose and glutathione; and the cell protection agent comprises polyethylene glycol 400. The electroporation transfection protection fluid can significantly reduce the deformity rate and mortality rate of abalone cells after electroporation technology, improve the gene transfection and gene editing efficiency, and reduce the cost of gene editing experiments, thereby laying a foundation for accelerating marine biological research and promoting environmental protection, species conservation and aquaculture.

The invention belongs to the technical field of biological gene modification, and particularly relates to transgenic moss, an ozone monitoring biosensor and a preparation method and application of the transgenic moss and the ozone monitoring biosensor. The transgenic moss is obtained by transfecting a target gene into moss cells, the target gene comprises a ZAT12 promoter, and the sequence of the gene ZAT12 is as shown in SEQ ID NO: 1. The transgenic moss is transferred into an oxygen response element ZAT12 promoter, and an ozone specific signal of the transgenic moss can be actively enhanced, so that high-sensitivity (5ppb) detection of ozone can be realized.

The application provides a gene transfection device and a gene transfection method. The gene transfection device comprises an ultrasonic transfection chip for generating ultrasonic waves and a reaction channel in contact with the ultrasonic transfection chip to receive the ultrasonic waves, and the reaction channel is used for loading a cell solution. The device provides a novel, simple, high-throughput, fast, safe and efficient solution for cell transfection, has the characteristics of wide application range, simple operation, good repeatability, strong stability and the like, and can flexibly realize efficient transfection of adherent cells and suspension cells, and has good applicability to various difficult-to-transfect cells such as stem cells, primary cells and tumor cells.

The invention relates to the technical field of plant genetic engineering, in particular to a mutant OsPBL8 gene, a preparation method thereof and application of the mutant OsPBL8 gene in enhancing rice disease resistance. The mutant OsPBL8 gene comprises an OsPBL8 gene mutant promoter full length and an OsPBL8 gene coding sequence full length, and the nucleotide sequence of the mutant OsPBL8 gene is shown as SEQ ID No: 5. The mutant OsPBL8 gene sequence is loaded on a pZ469 (pZCB11-1300-3xMyc-GFP) vector subjected to BamHI single enzyme digestion, then agrobacterium tumefaciens is transferred, and the gene is transfected into an initial rice plant by an agrobacterium tumefaciens-mediated transformation method to form transgenic rice. By modifying the promoter of the OsPBL8 gene, the disease resistance of the rice can be remarkably improved, so that a mutant OsPBL8 gene overexpression plant still has excellent rice blast resistance in an environment with a relatively large day and night temperature difference.

The invention relates to the technical field of biological materials, in particular to lipid nanoparticles based on modified natural saponin, a preparation method and application of the lipid nanoparticles, and the modified natural saponin comprises natural saponin modified by metal complexes and / or natural saponin modified by targeting molecules. The natural saponin is selected from at least one of glycyrrhizic acid, aescin, soyasaponin, ginsenoside, ursolic acid and oleanolic acid. The modified natural saponin can effectively simulate the structural function of cholesterol compounds in lipid nanoparticles, can overcome the problems of unstable physicochemical properties of cholesterol, potential safety hazards of sources and the like, and can significantly improve the gene transfection and silencing efficiency of nucleic acid drugs as a drug carrier. And a new solution is provided for gene therapy and vaccine development. The preparation method of the lipid nanoparticles based on the modified natural saponin is mild in reaction condition, simple and convenient to operate, easy to control and beneficial to industrial production.

The invention discloses a chemotherapeutic drug-resistant tumor cell model as well as a construction method and application thereof. According to the construction method of the chemotherapeutic drug resistance tumor cell model, the chemotherapeutic drug resistance tumor cell model is obtained by transfecting a gene for coding a BAD mutant BADS99D to tumor cells, the BAD mutant BADS99D is a protein obtained by substituting the 99th serine Ser of a BAD protein with aspartic acid Asp, and the amino acid sequence is as shown in SEQ ID NO.1. The invention further discloses a construction method of the chemotherapeutic drug resistance tumor cell model. The phosphorylation state of a BAD gene is simulated through point mutation of a BADS99 site by a transgenic technology, so that chemotherapy drug-resistant tumor cells are constructed, and the regulation and control effects of phosphorylated BAD on the biological characteristics of drug resistance, proliferation, migration, invasion, self-renewal and the like of cancer cells are analyzed on the molecular level.

Disclosed herein are sonoporogenic methods that allow for repeating sonoporogenic gene therapy treatments in a safe and efficient manner to improve nucleic acid delivery and expression in target cells. The methods include a combination for multiplex application of ultrasound in combination with application of an acoustically active agent and a nucleic acid payload to increase delivery of the nucleic acid payload to target cells in the subject, thereby increasing gene transfection and expression, and in some cases, in some cases, the delivery of the nucleic acid payload to the target cells in the subject. A process to increase the persistence of gene expression following treatment by sonoporosis-based gene therapy. The methods disclosed herein can include providing ultrasound energy to a plurality of locations in a target tissue comprising the target cells, and can apply ultrasound energy during a plurality of repeatable treatment procedures.

The invention discloses a method and system for screening and evaluating active pharmaceutical ingredients based on in-vitro culture, and relates to the technical field of enzyme activity detection.The method comprises the steps that a cell population expressing a target metabolic enzyme is constructed through gene transfection, and a fluorescently-labeled substrate detection system is established; processing the cell population by adopting a concentration gradient to obtain dynamic response data, and analyzing an enzyme activity change rate to recognize a drug action time point; an enzyme activity recovery process is monitored after the drug is removed, a reversibility value is calculated through an activity recovery curve, and a safety value is determined by combining a ratio of a maximum tolerance concentration value to a minimum effective concentration value; and evaluating the drug activity based on the reversibility value and the safety value, and screening to obtain candidate drugs. According to the method, systematic evaluation and efficient screening of the active pharmaceutical ingredients can be realized.

Provides is engineered cells, including induced pluripotent stem cells (iPSCs) and hematopoietic cells derived therefrom, pharmaceutical compositions comprising said cells, and methods of use of same, wherein one or more genes are transfected into said cells, such that bone marrow homing is enhanced.

The invention discloses an activated release platelet drug delivery system as well as a preparation method and application thereof. The system comprises platelets and a gene transfection nano-composite entrapped in cytoplasm of the platelets, and the composite is formed by electrostatic self-assembly of reactive oxygen species (ROS) responsive cationic polymers and nucleic acid drugs. The preparation method comprises three main steps of platelet separation and purification, ROS responsive gene transfection nanocomposite preparation and platelet drug loading. The platelet is used as a natural carrier, gene drugs can be effectively entrapped and protected, fixed-point activated release of the drugs is achieved in a high-ROS microenvironment of an injured part, and excellent targeting, biocompatibility and controllable release characteristics are achieved. The system is simple and convenient in preparation process and mild in condition, is suitable for gene therapy of tissue injury diseases such as spinal cord injury, and has important clinical application value.

The invention discloses preparation and application of a hyperbranched polymer with high gene transfection capacity, and belongs to the technical field of chemical synthesis and modification. The preparation method of the hyperbranched polymer with high gene transfection capacity comprises the following steps: adding an acrylate compound and organic amine into an organic solvent, dissolving, carrying out addition reaction, cooling, adding an end-capping reagent, and continuously reacting to obtain the hyperbranched polymer. The hyperbranched polymer prepared by the invention can solve the problem that the transfection efficiency is low after the existing precursor is polymerized, the transfection capacity of nucleic acid drugs in cells is remarkably improved while the biological safety is ensured, and the hyperbranched polymer is expected to replace viral vectors to be used as mainstream nucleic acid drug delivery vectors in the market.

The utility model discloses an ultrasound mediation gene transfection device which comprises a transfection box, a supporting partition plate is fixedly connected to the lower end of an inner cavity of the transfection box, an ultrasonic generator is fixedly installed at the middle end of the bottom of the supporting partition plate, a through hole is formed in the surface of the supporting partition plate, and a transfection container is movably connected to the top of the supporting partition plate. By arranging the transfection box and the supporting partition plate, a transfection container containing cell sap can be supported in the gene transfection process, and by arranging the driving motor, the first screw rod, the fixed frame, the movable frame, the movable frame, the energy converter, the through hole, the double-shaft motor, the second screw rod, the movable plate, the push rod and the ultrasonic generator, the cell sap can be supported in the transfection process. The purpose of ultrasound-mediated gene transfection is achieved, and corresponding operation can be automatically performed on cell sap in the transfection container one by one in the gene transfection process, so that the transfection operation quality is effectively ensured.

A cell mixture for the patient-specific treatment of solid tumors comprises carrier cells with a special loading. Immunocompetent autologous cells and / or genetically transfected cells are used, loaded with a combination of immunocompetent components and / or theranostic nanoparticles. The target tissue is derived from a tumor and includes malignant tumor cells and deactivated intratumoral cells, which are subsequently separated. Liposomes are loaded with a patient- and tumor-specific combination of immunocompetent components, namely at least one stable isotope with a high affinity for (epi-)thermal neutrons and / or a β- +The process involves the use of "decay" isotopes and / or a sensitizer to perform photodynamic therapy. Additionally, the cells are loaded with theranostic nanoparticles, which also allow for monitoring of the cells and charges involved. The carrier cells are gently loaded with the loaded liposomes via passive fusion. The loading rate of the carrier cells is significantly higher than with conventional methods, and at the same time, the carrier cells are highly specific to the individual tumor of the patient.

The invention discloses a nucleic acid delivery system and a preparation method and application thereof, and belongs to the technical field of biological materials. According to the invention, a benzenesulfonamide functional group is introduced into a GSs molecular skeleton, such that a novel benzenesulfonamide functionalized gemini surfactant (NGS) is constructed. Wherein benzenesulfonamide is a specific endoplasmic reticulum targeting group, and can realize endoplasmic reticulum targeting enrichment through specific binding with a sulfonylurea receptor on an endoplasmic reticulum membrane, so that the NGS gene delivery vector is distributed towards the endoplasmic reticulum. After the NGS and the pDNA form a compound, the compound enters cells through cavette protein mediated endocytosis, then targets the endoplasmic reticulum and releases the pDNA, the pDNA is delivered into the cell nucleus by utilizing the close relation between the endoplasmic reticulum and the nuclear membrane, the gene transfection efficiency is finally improved, meanwhile, the biocompatibility is further improved by introducing the gamma-polyglutamic acid (gamma-PGA) with negative charges, and the application prospect is wide.

An effective gene therapy technique that can transfect a gene to a cell in an organ or a tissue in a subject in a safe, effective, and durable manner is disclosed herein. The methods disclosed herein can induce expression of a nucleic acid payload in target cell(s) of a mammalian subject by administering to the subject a linear double stranded DNA (dsDNA) construct covalently closed at each of its ends by DNA loops, the linear dsDNA construct comprising the nucleic acid payload; administering to the subject a plurality of microbubbles; and administering to the subject, in proximity to the target cell(s) an effective amount of an ultrasound energy, thereby inducing the formation of pores the target cell(s) by disrupting the plurality of microbubbles to result in introduction of the dsDNA into the target cell(s) and expression of the nucleic acid payload in the target cell(s).

The invention discloses an amino acid modified poly (2-hydroxypropyleneimine) gene vector and a preparation method thereof. The method comprises the following steps: adding EDC / NHS activated amino acid into a PHP solution, adjusting the pH value of a reaction mixture to 5.5 + / -0.5, and reacting at normal temperature to prepare the amino acid modified PHP gene vector. Compared with a traditional PEI material, the amino acid modified PHP gene vector disclosed by the invention is higher in gene transfection efficiency and stronger in transfection performance.

Accordingly, the present disclosure provides a population of genetically engineered mesenchymal stem cells (MSCs), comprising an expression vector comprising an Akt or HGF gene and a PD-L1 gene. Also provided is a method for synergistically increasing survival status and immunomodulatory ability of an MSC or enhancing proliferation of an MSC, comprising transfecting an MSC with an Akt or HGF gene and a PD-L1 gene and a method for preventing, ameliorating and / or treating an ischemia condition, enhancing neuroregeneration or reducing neuronal death, comprising administering an effective amount of a population of genetically engineered MSCs of the present disclosure to a subject in need thereof.

The subject invention is directed to a mixed cell composition to generate a therapeutic protein at a target site by providing a first population of mammalian cells transfected or transduced with a gene that is sought to be expressed, and a second population of mammalian cells that have not been transfected or transduced with the gene, wherein endogenously existing forms of the second population of mammalian cells are decreased at the target site, and wherein generation of the therapeutic protein by the first population of mammalian cells at the target site stimulates the second population cells to induce a therapeutic effect.

Provided are a UV light-responsive hyperbranched poly-β-amino ester having high-efficiency gene delivery ability and a preparation method and application thereof; said poly-β-amino ester uses 4-amino-1-butanol, 2-nitro-1, M-phthaloyl 3-diacrylate, trimethylolpropane triacrylate, and 1-(3-aminopropyl)-4-methylpiperazine as raw materials, is polymerized by means of the “A2+B3+C2” Michael addition method, causing it to have a hyperbranched structure. In comparison with a linear structure, the branched structure enhances the interaction between the polymer and the nucleic acid molecule, significantly improving gene condensation ability, while also increasing cellular uptake by means of enhancing the interaction with the cell membrane. The poly-β-amino ester has a UV-responsive group on the backbone chain; under UV light irradiation, the poly-β-amino ester can be rapidly degraded after endocytosis, and releases the encapsulated genes, and achieves efficient gene transfection and reduces material toxicity. The invention has good prospects for development in the field of biomedical materials, and particularly in gene delivery.

The function of enhancing stem cells is accomplished with a specific transfected human gene type into a stem cell of choice to allow for the regenerative potential of the selected stem cell to be greatly enhanced by combining both gene and stem cell therapies together. Specific genes for selected protein production are cultured with a stem cell of choice which then produces a supernatant composed of an exosome-gene complex. This exosome-gene complex can also be used as a therapeutic gene therapy on its own or in combination with a stem cell gene complex together for a specific cellular effect, organ or tissue of regenerative effect or general systemic result.

The present invention addresses the problem of avoiding an immune response caused by gene transfection. The present inventors have found, after intensive studies to solve the above problem, that it is possible to avoid apoptosis due to innate immunity and increase the number of transgenic cells by suppressing innate immunity in cells into which a gene is transfected. On the basis of this finding, further studies have been conducted to complete the present invention.

A novel cationic lipid has a structure as represented by general formula (1) and specifically relates to a nitrogen-branched cationic lipid, and a liposome containing the cationic lipid, and a nucleic-acid pharmaceutical composition containing the liposome, a preparation method and application thereof, wherein, the definition of each symbol in the formula (1) is as defined herein. The cationic liposome containing the cationic lipid as represented by formula (1) can improve the loading rate and transport efficiency of nucleic-acid drugs. The formulation of the cationic liposome nucleic-acid pharmaceutical composition has good cell compatibility and higher gene transfection capability, and can improve the treatment and / or prevention effects of nucleic-acid drugs.

The application discloses a polymer containing a chemotherapy drug SN38 and a macrocyclic polyamine, and a preparation method and application thereof, and belongs to the technical field of polymers.The polymer containing the chemotherapy drug SN38 and the macrocyclic polyamine can release the SN38 under esterase conditions; can effectively condense siRNA to form uniform spherical nanoparticles; the nanoparticles formed by compounding with auxiliary lipids (DSPE-PEG-iRGD) have excellent tumor targeting and selectivity; under hypoxic conditions, the gene transfection capacity can be improved through a photochemical internalization effect to promote lysosome escape, and the knockdown rate of Bcl-2 protein is as high as 76%; the proliferation of HCT116 cells in the mouse body is significantly inhibited through delivery of Bcl-2 siRNA and photodynamic therapy, and the tumor inhibition rate is as high as 95%. The polymer can be used for preparing a non-viral gene carrier, as a photosensitizer, and for preparing a carrier for delivering the chemotherapy drug SN38 in tumor treatment.

The invention provides an establishment method and application of a flavidae gill filament cell line. The preservation number of the cell line is CGMCC (China General Microbiological Culture Collection Center) No.46720. The flavidae gill filament cell line established by the invention has good hereditary stability, and continuous passage of more than 100 generations is realized. A gene transfection experiment verifies the efficient transfer and expression ability of the cell line to exogenous genes, can provide abundant cell resources for the research of gene functions of the Flavonus flavipes and the related research of molecular biology and the like, and has a wide application prospect.