78 results about "Cellular engineering" patented technology

The present application relates to the field of plant tissue culture technology, and particularly discloses a lily stem cell abundance ex vivo regulation method. The lily bulb is used as the starting material, stem cell magnitude amplification and regulation culture can produce lily tissue organ materials with high stem cell abundance. The present application provides sterile lily materials with high stem cell abundance, which has the characteristics of strong regenerative ability, high proliferation rate, etc., is the basis for protoplast preparation and high-efficiency regeneration, and is suitable for different lily varieties, and provides technical support for cell engineering breeding, seedling scale breeding, etc.

The invention relates to the field of cell engineering and gene engineering, in particular to a method for enhancing the multiplication capacity of umbilical cord mesenchymal stem cells and application. The expression or activity of the STN1 gene in the umbilical cord mesenchymal stem cells is reduced through an accelerant, and the umbilical cord mesenchymal stem cells with enhanced multiplication capacity are obtained. By knocking down the hUC-MSCs of STN1, the proliferation speed of the hUC-MSCs is higher, DNA replication is more active, the clone formation ability is higher, and the aging proportion of the hUC-MSCs can be remarkably reduced.

The invention relates to the related technical field of molecular biology and cell engineering, in particular to an FLP-IN-PIEC cell line and a construction method and application thereof, and the construction method comprises the following steps: step 1, constructing CRISPR / Cas9 site-specific cleavage plasmid containing a pig HIPP11 safety site sequence; step 2, constructing a donor plasmid Donor-FRT containing a homologous arm of a safety site of the HIPP11; 3, jointly transfecting plasmids in the step 1 and the step 2 into a porcine hip artery endothelial cell line (PIEC) to obtain an FLP-IN-PIEC cell line for expressing red fluorescence; and 4, under the action of Flp recombinase, transfecting an exogenous gene donor plasmid containing an FRT site to the FLP-IN-PIEC cell line obtained in the step 3, and driving the expression of a target gene through an optimized strong promoter to realize fixed-point transfer and efficient expression of the exogenous gene in the pig (PIEC) cell line. Different from an existing Flp-In system depending on a random integration strategy, the construction method disclosed by the invention emphasizes fixed-point integration and promoter optimization, and improves the homogeneity, stability and screening efficiency of exogenous gene expression.

The present disclosure relates to immunotherapy. In more specific embodiments, the present disclosure provides systems, compositions, methods and uses of viral vectors comprising nucleic acid sequence of interest that encodes at least one therapeutic product, and a nucleic acid sequence encoding at least one nuclease, for in vivo targeted insertion of the nucleic acid sequence of interest into a target locus within at least one cell of the T lineage.

This invention discloses an immune cell that secretes type 2 cytokines under hypoxic conditions and its applications, belonging to the fields of genetic engineering and cell engineering. The invention first constructs a nucleotide fragment encoding hypoxia-induced type 2 cytokines, including a signal peptide gene sequence, a CAR molecule nucleotide sequence targeting CD19 and CD22, and an IL-4 or IL-10 nucleotide sequence containing multiple HRE motifs of a hypoxia-inducible promoter. This nucleotide fragment is delivered to immune cells for stable expression, yielding an immune cell that secretes type 2 cytokines under hypoxic conditions. This immune cell secretes IL-4 or IL-10 only under hypoxic conditions, thereby leveraging the metabolic reprogramming effect of IL-4 or IL-10 on immune cells to enhance their activity under hypoxic conditions, thus achieving better long-term anti-tumor effects.

The application belongs to the technical field of animal cell engineering, and particularly relates to a pigeon fibroblast cell strain X4 and application thereof. A pigeon fibroblast cell strain is provided, which is named pigeon fibroblast cell X4, and the preservation number is CGMCC No. 46754. The pigeon fibroblast cell strain can be used for virus isolation or amplification, virus pathogenic mechanism research, and development of antiviral drugs and vaccines, and has a good application prospect in pigeon disease research.

The invention provides a larimichthys crocea head kidney macrophage line as well as a construction method and application thereof, belongs to the technical field of cell engineering, and particularly provides the larimichthys crocea head kidney macrophage line which is named as LYC-hK and has the preservation number of CCTCC NO: C2025234. The larimichthys crocea head kidney macrophage line is constructed through the steps S1-S6, and the construction method of the larimichthys crocea head kidney macrophage line is applied to the field of cell engineering. The larimichthys crocea head kidney macrophage line provided by the invention can be used for research on pathogenic infection and pathogenic mechanism of larimichthys crocea, research on anti-infection immune response of larimichthys crocea and research on exogenous gene expression.

The invention belongs to the cross technical field of cell engineering, nano-drugs and biological manufacturing, and particularly discloses a drug-loaded vesicle based on denucleated cells as well as a preparation method and application of the drug-loaded vesicle. According to the invention, a cell suspension and a cytoskeleton relaxant are co-incubated to relax an actin skeleton and weaken nucleoplasm connection; then loading the treated cell suspension on a multi-layer discontinuous density gradient centrifugal medium, realizing physical separation of cell nucleuses and cytoplasm through high-speed centrifugation according to buoyancy density difference, and collecting components of a specific interface to obtain high-purity denucleated cells with a complete membrane structure; then co-incubating the denucleated cells and ROS response type lipidosome (co-carrying therapeutic siRNA and a sound-sensitive agent Ce6) prepared in advance, so that the lipidosome is wrapped or anchored by a denucleated cell membrane; finally, the composite system is subjected to extrusion treatment through a microporous membrane, the drug-loaded vesicles uniform in particle size and stable in structure are obtained, and the drug-loaded vesicles are suitable for various application scenes such as anti-tumor treatment and RNA vaccine delivery.

The application provides a method for constructing an immortalized bovine nasopharyngeal tonsil epithelial cell line and application, and belongs to the technical field of cell engineering. The healthy yellow bovine nasopharyngeal tonsil is used as a material, and the primary epithelial cells are separated by enzyme digestion. The 3rd to 4th generation cells are taken, infected with SV40 T lentivirus (MOI=80), and screened by 1.5 μg / mL puromycin to obtain a stable immortalized cell line. The cell line is continuously passaged to the 50th generation and still stably expresses SV40LT protein and epithelial specific markers CD326, CD324 and KRT8, maintains a typical pavement stone-like morphology, normal proliferation and no malignant transformation. Infection experiments show that the cell is highly sensitive to FMDV, supports efficient replication, and the virus titer can reach 10 7.5 TCID 50 / mL. The application overcomes the defects of the existing immortalized bovine nasopharyngeal tonsil epithelial cell model, such as limited source, large batch difference, low sensitivity and poor persistence, and provides a stable and reliable in vitro model for the research of foot-and-mouth disease prevention and control related mechanisms, drug screening and vaccine development.

The invention belongs to the technical field of animal reproductive biology and cell engineering, and particularly relates to a fluorescence labeling expression system for visualizing an oocyte nucleus maturation process and application of the fluorescence labeling expression system. The fluorescence labeling expression system comprises an eukaryotic expression vector pVenus-LMNB1; the eukaryotic expression vector pVenus-LMNB1 comprises a nucleotide sequence for coding the fusion protein Venus-LMNB1. According to the invention, a nuclear fiber layer protein stably expressed on a nuclear membrane is selected as a research object, a eukaryotic expression vector pVenus-LMNB1 is constructed by using a molecular cloning technology, an LMNB1 gene containing a green fluorescent label Venus is overexpressed in a porcine oocyte by using a microinjection technology, fluorescent protein labeling of a structural protein on the nuclear membrane is utilized, and the expression vector pVenus-LMNB1 is constructed. The expression positioning condition of the LMNB1 in the maturation process of the porcine oocytes is observed, and a powerful tool is provided for development of in-vitro culture related researches of the porcine oocytes.

The present invention relates in general to a platform to selectively eliminate unwanted cells from a population of cells. The invention relates to self-amplifying RNA constructs for cell purification preferably during differentiation and cell engineering.

The present invention provides a cell suitable for gene engineering, cell engineering and cellular medicine, and a method for producing the same. The present invention provides a technique for removing repetitive sequences present in a specific gene region in two or more alleles, thereby facilitating gene targeting or sequencing in the region.

The invention belongs to the technical field of cell engineering and genetic engineering, and particularly relates to application of an FBXO32 gene in regulation and control of skeletal muscle satellite cell proliferation and feed utilization rate. According to overexpression of the FBXO32 gene in skeletal muscle satellite cells, it is found that the mRNA expression quantity of skeletal muscle satellite cell proliferation marker genes KI67 and CCND1 is remarkably increased, and the protein expression level is also remarkably improved. CCK-8 and EdU detection results also show that the FBXO32 gene or the encoded protein thereof also has a promotion effect on skeletal muscle satellite cell proliferation. The overexpressed FBXO32 gene or the encoded protein thereof can promote the proliferation of skeletal muscle satellite cells, and lays a foundation for improving meat traits of livestock and poultry, improving the feed utilization rate of the livestock and poultry and improving the meat traits of the livestock and poultry.

The patent discloses a method for quickly establishing a Cabernet Sauvignon grape pericarp callus regeneration system, and belongs to the technical field of cell engineering. The method takes the pericarp of Cabernet Sauvignon grape as an explant, realizes efficient induction and rapid regeneration of callus through three key steps of explant disinfection treatment, primary callus induction culture, callus proliferation and subculture. The method optimizes the culture medium formula, adds specific concentrations of 6-BA, KT, IAA, NAA and 2,4-D in the induction medium, and significantly improves the induction rate and growth rate of callus. Experimental results show that the stable callus regeneration system can be obtained in 40-50 days by using the method, the callus induction rate can reach more than 90%, and the survival rate of regenerated plants is high. The patent has the advantages of simple operation, good repeatability and short cycle, and provides an effective technical means for genetic improvement, gene function research and rapid propagation of wine grapes, and has important theoretical value and application prospect.

The invention provides a drug combination composition for in-vivo target cell engineering modification, and target cells do not include myeloid cells. The drug combination composition comprises a first drug and a second drug, wherein the active ingredient of the first drug is a substance capable of saturating, inhibiting or eliminating phagocytic ability of in-vivo myeloid cells, and the active ingredient of the second drug is an in-vivo target cell capable of being engineered. Through combined application of the myeloid cell inhibition drug and the cell engineering modification drug targeting the target cells, endocytosis of the myeloid cells to the target drug is reduced or eliminated before engineering modification of the target cells, so that the off-target effect is reduced, the lymphatic organ targeting property can be improved, and the treatment effect of the myeloid cells is improved. The toxic and side effects of cell engineering modification drugs on the liver are reduced, and the method has important significance on in-vivo (in-situ) lymphocyte engineering modification (including gene editing) or immunotherapy (such as in-vivo CAR T cell therapy).

The application belongs to the technical field of cell engineering, and particularly relates to a bispecific antibody targeting CD22 and CD19, a chimeric antigen receptor thereof and application. In the bispecific antibody, a CD22 light chain and a CD22 heavy chain are connected through a linker 4 to form a CD22 single-chain antibody, and a CD19 light chain and a CD19 heavy chain are connected through a linker 4 to form a CD19 single-chain antibody, which comprises one of the following structures: (1) the CD22 single-chain antibody and the CD19 single-chain antibody are connected through a linker 1 or a liner 2; (2) the two ends of the CD19 single-chain antibody are connected with the CD22 light chain and the CD22 heavy chain through a linker 3, respectively. The chimeric antigen receptor provided by the application can simultaneously target two different tumor antigens, improve the killing of tumor cells, reduce the probability of immune escape, and reduce the tumor recurrence rate after CAR-T treatment.

This invention belongs to the field of cell engineering technology, specifically relating to an engineered mitochondrial nanoshield, its preparation method, and its applications. The engineered mitochondrial nanoshield can resist damage from high concentrations of calcium ions in the extracellular environment, and also possesses BMMSCs targeting ability and ROS-responsive characteristics. After intravenous injection, the engineered mitochondrial nanoshield can selectively target BMMSCs to alleviate cellular senescence and increase bone mass in aged mice.

The invention provides pichia pastoris for producing a honey truffle sweetening agent as well as a construction method and application thereof, and relates to the technical field of microorganisms. According to the invention, high-efficiency expression of a natural sweet protein honey truffle sweetening agent is realized in pichia pastoris cell engineering, a screened signal peptide C terminal is connected with a Pro region of an alpha mating factor to obtain a fusion signal peptide, and heterologous expression of a honey truffle sweetening agent coding gene is realized through the fusion signal peptide. The pichia pastoris capable of efficiently secreting and expressing the target sweetening agent in an extracellular mode is successfully constructed, the pichia pastoris can be used for efficiently producing the honey truffle sweetening agent, the shake flask yield reaches 44 mg / L, and an efficient and feasible new strategy is provided for large-scale microbial production of the honey truffle sweetening agent.

This invention relates to the field of cell engineering technology, and more particularly to a method for preparing immunomodulatory stem cells for treating graft-versus-host disease (GVHD). The invention provides a method for preparing immunomodulatory stem cells for treating GVHD, comprising the following steps: (1) washing human fat aspirate and digesting it with collagenase to obtain monocytes; (2) inoculating the monocytes obtained in step (1) into differentiation medium and culturing for 2–3 days, then continuing to culture the adherent cells for 18–22 days to obtain mesenchymal stem cells; (3) inoculating the mesenchymal stem cells obtained in step (2) into induction medium for induction culture, and after screening, obtaining immunomodulatory stem cells. The preparation method provided by this invention solves the technical problems of unstable sources, functional decline, and poor therapeutic effects of immunomodulatory stem cells in the prior art, providing a new, efficient, safe, and standardized solution for the treatment of GVHD.

Conventional CAR-T cell therapies have shown a significant success in the treatment of hematological cancers and lymphoma. However, this technique still has several problems, including cancer recurrence. Herein, a scalable technical platform is described to produce synthetic viscoelastic activated cells (SynVAC) with programmable mechanical and chemical activity as artificial antigen presenting cells (aAPC). The disclosure presented herein shows that the viscoelastic properties of the described SynVAC have great beneficial effects on expansion of T cells. For example, SynVAC exhibits robust improvements in T cell expansion, T memory stem cell (TMSC) formation, chimeric antigen receptor (CAR) transduction efficiency, tumor killing efficiency, and long-term in vivo persistence of CAR-T cells compared to conventional rigid or elastic microspheres.

The invention discloses a method for establishing an efficient organ in-vitro regeneration system of improved hybrid liriodendron species, and belongs to the technical field of plant cell engineering. The method comprises the following steps: obtaining a hybrid liriodendron chinense improved seed somatic embryo regenerated plant which is subjected to illumination culture for 60 days, taking a stem segment or a stem tip as an explant for in-vitro bud induction, screening out a culture medium formula which is most suitable for the stem segment and the stem tip, and vertically placing the stem segment; carrying out proliferation and elongation culture on the induced in-vitro buds under specific conditions, and screening out the most suitable proliferation culture medium formula and elongation culture medium formula; carrying out rooting culture on the elongated buds, and screening out an optimal rooting culture medium formula; finally, seedling hardening and transplanting are conducted, the seedling hardening method is that bottles are sealed for 3 days, the bottles are opened for 1 day, and a transplanting matrix is prepared from humus nutrient soil and perlite according to the ratio of 7: 4. The method is high in regeneration efficiency, simple and convenient to operate and capable of meeting large-scale production requirements.

The invention belongs to the technical field of animal cell engineering, and particularly relates to a pigeon fibroblast strain X4 and application thereof. The invention provides a pigeon fibroblast strain which is named as pigeon fibroblast X4 and has a preservation number of CGMCC (China General Microbiological Culture Collection Center) No. 46754. The pigeon fibroblast cell strain can be used for separation or amplification of viruses, research of pathogenic mechanisms of the viruses and development of antiviral drugs and vaccines, and has a good application prospect in pigeon disease research.

The present disclosure generally relates to engineered adeno-associated virus and their uses in T cell engineering. Also disclosed are engineered viral capsid polypeptides, vectors, cells, and kits, related to the engineered adeno-associated virus, and methods related to the uses thereof.

The application provides a mesenchymal stem cell cryopreservation solution and a preparation method thereof, and belongs to the technical field of cell engineering. The cryopreservation solution takes diluted DMEM as a basic culture medium, and introduces trehalose, L-proline, polyethylene glycol, sodium carboxymethyl cellulose, sodium glycine, sericin, glycerol, resveratrol and other components as additives. The cryopreservation solution formula adopts a DMSO-free and serum-free scheme, uses sodium glycine, sericin and glycerol as a composite protective agent, can effectively alleviate ice crystal damage, maintain osmotic pressure balance and protect cell membranes and protein structures, and has an effect superior to DMSO and glycerol and no cytotoxicity. In addition, the application uses resveratrol, coenzyme and astragaloside and other components as growth factors to maintain the activity of mesenchymal stem cells in a frozen state. After umbilical cord mesenchymal stem cells are cryopreserved for 2 years according to the application, the survival rate of the umbilical cord mesenchymal stem cells can reach 99.83% at most, the cell viability OD value reaches 0.96, and the cells are observed to have good adhesion and growth conditions under a microscope. △ The survival rate of the umbilical cord mesenchymal stem cells can reach 99.83% at most, the cell viability OD value reaches 0.96, and the cells are observed to have good adhesion and growth conditions under a microscope.

The invention discloses a model building method based on animal endometrial epithelium organoid and application of the model building method, and relates to the technical field of cell engineering and organoid. According to the method, healthy cow endometrial tissue serves as a raw material, a model is built through the steps of pretreatment, enzymolysis, cell cleaning and screening, seed gum culture, passage, cryopreservation resuscitation and the like, and the model is built through Hamp; e, verifying morphological integrity through dyeing, and detecting E-cadherin expression through IHC dyeing to confirm cell specificity. The animal organ model restores the in-vivo three-dimensional microenvironment and species specificity, overcomes the limitation of traditional two-dimensional culture and mouse models, can accurately simulate the pathological process of endometritis, is applied to research on the pathogenesis of endometritis of dairy cows, can also efficiently screen anti-inflammatory drugs and evaluate the drug effect, and has a wide application prospect. A reliable in-vitro tool is provided for disease prevention and treatment, and healthy development of the dairy cow breeding industry is assisted.

The invention is applicable to the technical field of cell engineering, and provides a preparation method and application of a multifunctional glycoprotein photo-crosslinking agent, and according to the method, a biotin group type crosslinking agent containing a trifluorobis (acridine) photo-reaction active group, an oxamine sialic acid specific recognition group, a chain length-adjustable polyethylene glycol chain and a disulfide bond cleavable is prepared. The prepared multifunctional glycoprotein photo-crosslinking agent can realize efficient photo-induced protein crosslinking, and the structural design of the multifunctional glycoprotein photo-crosslinking agent is beneficial to subsequent protein decrosslinking and protein compound identification. By utilizing a trifluoro-bis-acridine photoreaction active group, the trifluoro-bis-acridine photo-crosslinking polymer has excellent photo-crosslinking activity; specific recognition and reaction on a sialic acid-containing sugar chain are realized by using an oxygen amine group; a polyethylene glycol chain with adjustable chain length is utilized to improve the solubility and biocompatibility, and cytoplasm pollution is avoided as far as possible; by utilizing biotin groups of disulfide bonds, efficient enrichment is realized, and the biotin groups can be selectively cut under a reduction condition.

The present application relates to a kind of enrichment method of stem cell high purity positive cell group, belong to the field of biotechnology and cell engineering technology.It aims to solve the technical problems such as the cell damage of existing screening technology, low screening efficiency, easy to affect stem cell pluripotency and genome stability, etc.The present application forms 23-cell small mass after H9 human embryonic stem cells infected by lentivirus are digested gently, after gradient dilution static culture, high expression positive clone is screened under fluorescence microscope, and target cell group is directly enriched by visual mechanical scraping.The present application does not change cell genome, and is high in safety;Small mass inoculation significantly improves cell survival and cloning efficiency at low density;Selective scraping operation is accurate, fast and low in cost;The proportion of reporter gene (EYFP) positive cells in the obtained positive cell group can reach 93.2%, and good pluripotency and function are maintained, suitable for efficient enrichment of stem cells and immune cells.

The invention discloses a preparation method and application of a novel macrophage for treating myocardial infarction, and belongs to the fields of cell engineering, nanotechnology, material science, immunology and the like. According to the invention, bone marrow-derived macrophages are induced to be polarized to M2 type and are subjected to magnetic modification, so that the macrophages are endowed with the capability of efficiently expressing vascular endothelial growth factors, Mag M2-M-VEGF cells are obtained, and proliferation of endothelial cells and formation of collateral vessels are promoted through local high expression of VEGF at a myocardial infarction part, so that local ischemia injury is relieved. According to the invention, precise targeted delivery of cells is realized, local inflammatory response is inhibited, and blood perfusion in an infarction area is improved. The medicine is endowed with magnetic targeting capacity by internalizing Fe3O4 NPs and is accurately enriched in myocardial infarction tissue under the guidance of a magnetic field, M2 type polarization inhibits pro-inflammatory reaction by reducing pro-inflammatory factor release and releasing a large amount of anti-inflammatory factors, and meanwhile, VEGF is specifically secreted, angiogenesis of an infarction part is promoted, and the capillary density of an infarction region is effectively increased.

The invention belongs to the field of cell engineering seedling culture of algae, and particularly relates to a cell engineering seedling culture method of gracilaria lemaneiformis. The gracilaria lemaneiformis frond is constructed into a miniaturized form, and high-density intensive culture is carried out by utilizing a photobioreactor. According to the method, the microminiaturized asparagus frond obtained by a cell engineering means can be subjected to vegetative propagation, efficient propagation is carried out by utilizing the algae photobioreactor, and the microminiaturized asparagus frond is further cultured into seedlings for artificial cultivation.