44 results about "Mice transgenic" patented technology

This invention discloses a method for constructing a biological model of tertiary lymphoid structures, which can rapidly and efficiently induce the generation of in situ TLS, without relying on specific tissues or organs, and without the need for complex and expensive preparation processes such as cell editing and transgenic mice. It can generate stable, mature, and long-lasting TLS structures within 2 to 3 weeks, and has the characteristics of short cycle, low cost, simple operation, reproducibility, safety and effectiveness.

The invention belongs to the technical field of biological genetic engineering, and particularly relates to a mouse model construction scheme capable of reproducing a key phenotype of hemophagocytic syndrome, which comprises the following steps: inserting a loxP-STOP-loxP-LMP1 conditional expression cassette into a Rosa26 safe port site in a fixed point manner to obtain an Lmp1cKI mouse strain; and hybridizing the mouse with an Ncr1-Cre transgenic mouse of NK cell specific expression Cre recombinase. In a double transgenic offspring (Lmp1Ncr1-Cre), STOP fragments are cut off by Cre enzyme, and continuous high expression of EB virus LMP1 protein in NK cells is driven. The model does not need chemical induction or immune intervention, and typical hemophagocytic syndrome phenotypes, namely serum IL-6, TNF-alpha and IFN-gamma, spontaneously appear within 7-9 days after young rats are born; the body weight is reduced and the survival rate is obviously reduced; the proportion of lymphocytes is sharply reduced; and NK cells are depleted. The invention provides a stable and reliable tool for hemophagocytic syndrome pathogenesis research, drug screening and treatment evaluation.

The application relates to a construction method of a Mettl3 transgenic mouse model with specific knockout, enriches the understanding of the function and action mechanism of Mettl3 in mature pancreatic islet beta cells, and provides a new thought and target for the pathogenesis and treatment of diabetes.

The present disclosure provides transgenic mice for use as mouse models, and methods related thereto. The transgenic mice described herein comprise a humanized mouse telomerase (hmTert) gene included in a background mouse germline that provides shortened telomeres capable of reproducing humans and various human disease states.

The invention belongs to the technical field of GPCR drug research and development, and discloses a ratio type cAMP fluorescent probe RaCamp and a construction method of a transgenic mouse, the probe RaCamp is formed by series fusion of G-Flamp2 fluorescent protein and mCherry fluorescent protein through a flexible linker, and the preparation method of the probe RaCamp comprises the following steps: S1, obtaining an mCherry fragment; s2, a G-Flamp2 linear carrier is obtained; s3, homologous recombination; s4, converting and screening; the construction method of the transgenic mouse comprises the following steps: T1, designing a carrier; t2, microinjection of fertilized eggs; t3, embryo transplantation and reproduction; the invention has the beneficial effects that mCherry is added as an internal reference protein on the premise of keeping the sensitivity of the original G-Flamp2 probe, and the selected connecting peptide (GGGGS) 2 enables the expression quantity of the two proteins in cells to be accurately controlled to be 1: 1, so that the problem of background interference in cell observation of in-vivo imaging is well solved. And the sequence is inserted into a Rosa26-LSL box, so that the construction of a Rosa26-LSL-RaCamp mouse is realized.

The invention belongs to the technical field of disease model construction, and particularly relates to a construction method of a chronic myelogenous leukemia mouse model with conditional human EZH2 overexpression and RUNX1 knockout. According to the invention, a chronic myelogenous leukemia mouse transgenic mouse model with conditional human EZH2 overexpression and RUNX1 knockout is successfully constructed, the model is induced to be converted from a chronic stage to a sudden change stage, and particularly, the model is a transgenic mouse model which is positive in Lyz2-CreERT2 / EZH2 / RUNX1 and carries BCR-ABL and SCL-tTA. It is proved that a human EZH2 conditional overexpression and RUNX1 knockout chronic myelogenous leukemia mouse transgenic mouse model has feasibility and importance for research on conversion from CML CP to BC samples, and a molecular mechanism for conversion from chronic myelogenous leukemia to a sudden change stage is revealed for research. And a new animal model and a new research idea are provided for understanding of disease progression and development of a new treatment strategy.

The invention discloses a p16-OSKM transgenic mouse model and a reprogramming senescence cell construction method and application, and belongs to the technical field of cell biology and regenerative medicine. According to the method, a p16-OSKM hybrid mouse capable of specifically inducing an OSKM factor in a p16 high expression cell is obtained on the basis of hybridization of a p16-Ink4a-CreERT2 transgenic mouse, a Rosa26-CAG-LSL-rtTA3-IRES-mKate2 transgenic mouse and a Col1a1-TRE-OSKM-IRES-mCherry transgenic mouse. The selective activation of the OSKM factor in senescent cells with high expression of p16 can be realized by adding doxycycline into primary embryo fibroblasts separated from the mouse under an in-vitro culture condition, so that the senescent cells enter a cell cycle again and recover division and multiplication capacities. The technology provides an accurate model platform for basic research, and can be used for aging mechanism exploration, cell fate regulation and control, chronic disease treatment, development and screening of anti-aging drugs and the like.

The present invention provides a basis for using soluble fiber, particularly compositions comprising Agave fructans, to prevent or treat the accumulation of lipids, particularly ceramides in the liver and brain and lipofuscin in the liver, improving the overall integrity of both organs, with treatment being feasible to prevent fatty liver disease and also prevent Alzheimer's disease (AD). In the present invention, we analyze alterations in lipid metabolism in the liver and ceramide production in the brain of transgenic mice (APP / PS1, TG) for AD. Our results indicate that microvesicular lipid accumulation within hepatocytes is increased in TG mice compared to the control group. Furthermore, in the brains of TG mice, a greater accumulation of ceramides is observed in the somatosensory cortex and entorhinal cortex; in the CA1 region of the hippocampus and in the motor cortex, no significant increase was observed when compared to the control group.

This invention belongs to the field of tumor biology and drug screening technology, specifically disclosing a human EGFR mutation-driven mouse primary lung cancer cell line, its construction method, and its applications. The cell line, ZST-1, is a human EGFR (L858R / T790M) mutation-driven lung cancer cell line derived from mouse primary lung cancer. It is stable, capable of subcutaneous tumor formation in C57BL / 6 mice, and simultaneously expresses Luciferase and tdTomato reporter genes. Its construction method includes obtaining transgenic mice, virus-induced tumor formation, continuous in vivo passage in nude mice, and in vitro culture and screening steps. This cell line can be applied to in vitro screening and efficacy evaluation of human EGFR mutation-targeting drugs, research on EGFR-TKI resistance mechanisms, tumor bioluminescence imaging and fluorescence tracing, and in vivo tumorigenesis and efficacy experiments in an immune-intact C57BL / 6 background.

The invention discloses a non-human animal integrated with a double-derived VH subclass fusion gene as well as a construction method and application of the non-human animal. The method comprises the following steps: firstly, constructing a gene construct containing human IGHV1-69 and IGHV3-23 fragments, a specific spacer region, a functional element and a rat heavy chain 3'enhancer, and microinjecting the gene construct into a C57BL / 6J fertilized egg with an endogenous heavy chain and a kappa light chain knocked out to obtain a transgenic mouse; through identification, after a mouse endogenous antibody is inactivated, a human gene is stably integrated, a human IgM antibody is expressed and H5N1 HA immunization is carried out, the use frequency of IGHV1-69 is remarkably increased, the mouse produces a high-titer specific antibody, and serum has neutralizing activity. The mouse can generate a target bnAbs aiming at the HA stem and the RBS in parallel, so that the antibody screening efficiency is improved, and an in-vivo screening and evaluation platform for simulating human body fluid response is provided for research and development of influenza antibodies and vaccines.

The invention discloses a ganoderma lucidum preparation for treating Parkinson's disease and application thereof, relates to a medicine, and provides the ganoderma lucidum preparation for treating Parkinson's disease, the ganoderma lucidum preparation comprises ganoderma lucidum hawthorn spore powder and ganoderma lucidum spore oil, the weight ratio of the ganoderma lucidum hawthorn spore powder to the ganoderma lucidum spore oil is (1-2): (1-2), and the weight ratio of the ganoderma lucidum hawthorn spore powder to the ganoderma lucidum spore oil is (1-2): (1-2). In the anti-senile dementia process, it is known that ganoderma lucidum has the protection effect on nerve cell mitochondria, nerve cell axons, nerve cell short processes and the like of a central nervous system, and has the effect of promoting growth of nerve cells of the central nervous system, and the effect is opposite to the pathological development of Parkinson's disease.

The present invention provides non-human animals expressing human CFB protein or chimeric (e.g., humanized) CFB protein and methods for using the same. The present invention further provides a non-human animal genome, a humanized CFB gene, and cells, tissues, organs, or non-human animals containing the non-human animal genome and the humanized CFB gene. The non-human animals obtained in this application successfully express human CFB protein or humanized CFB protein, which can be cleaved by CFD, bind to C3b, and exert similar effects in the human body. Furthermore, the non-human animals obtained in this application have complete renal function, normal blood biochemical indicators, and, more remarkably, do not exhibit the potential diseases observed in other transgenic mice known in this art.

The invention discloses an application of a NanoLuc luciferase double-subunit complementary system in construction of an experimental animal model. The genome of the experimental animal model contains a NanoLuc enzyme large subunit LgBiT encoding gene and stably expresses LgBiT; correspondingly, a tested object used for invading the experimental animal model or connected into the experimental animal model is in fusion connection with NanoLuc luciferase small subunit HiBiT polypeptide serving as a label or a NanoLuc luciferase small subunit HiBiT coding gene serving as a reporter gene, and LgBiT and HiBiT are specifically combined in the experimental animal model and recombined to form the NanoLuc enzyme with catalytic activity. A strong bioluminescence signal is generated under the action of a furazine substrate, so that high-sensitivity, non-invasive and real-time dynamic imaging of in-vivo distribution, loading capacity and diffusion path of a detected object is realized. The invention also discloses a gene editing system for constructing the LgBiT transgenic mouse.

The invention discloses a method for tracing glioma infiltrated microglial cells, and relates to the technical field of biological cell detection. The invention discloses a method for tracing glioma infiltrated microglial cells. The method comprises the following steps: hybridizing to obtain a transgenic mouse for tracing the microglial cells, inducing the microglial cells to express fluorescent protein, constructing an in-situ glioma model, collecting the brain of the mouse, slicing and dyeing. A mouse used in the invention is a transgenic mouse which traces TMEM119 expression, a fluorescent protein tdTomato is labeled on a microglial cell specific marker TMEM119, in a mouse glioma model, the spatial distribution of microglial cells in glioma can be intuitively observed by judging whether tdTomato fluorescence exists or not, and monocyte-derived macrophages are distinguished by co-dyeing with Iba1.

The present invention provides a method for constructing a Map3k3I441M mutation knock-in transgenic mouse model and a use. The method comprises: replacing the region from exon 1 to part of intron 2 of a mouse with a mutant gene fragment, wherein the sequence of the mutant gene fragment is as shown in SED ID NO: 1 in the sequence listing; using a BCA clone as a template to amplify a homologous arm-containing mouse genome fragment as a targeting vector; co-injecting Cas9 and gRNA as well as the targeting vector into fertilized eggs; allowing the fertilized eggs to develop to obtain positive F0 generation samples; co-housing the positive F0 generation samples for breeding to obtain F1 generation samples; performing genotyping on the F1 generation samples to screen for Map3k3I441M samples; and mating the Map3k3I441M samples from the F1 generation samples with a Cdh5-CreERT tool sample to produce F2 generation samples. A model constructed by the construction method can accurately reflect the real situation in a mouse.

The invention belongs to the field of bioengineering, and particularly relates to a construction method and application of a transgenic mouse for screening tissue cell membrane protein in vivo. Aiming at the problems of low in-vivo research flux, unstable library, limited tissue applicability and the like of the traditional membrane protein gene, sgRNA library plasmids are constructed by designing sgRNA of a membrane protein target gene and a control gene; three-positive-mouse containing Cas9, UBC-CreERT2 and sgRNA libraries is obtained through embryo microinjection and hybridization, genotype identification, inducer regulation and control, disease modeling, multi-tissue sample treatment and NGS analysis technical systems are established in a matched mode, and membrane protein gene in-vivo high-throughput screening is achieved. The model can guarantee library stability and screening reliability, covers multi-tissue research scenes, links in-vitro screening and in-vivo verification and is suitable for membrane protein function analysis and disease target mining, the technical process can be popularized to other gene families, and systematic biological research is promoted.

The invention discloses a construction method and application of a sperm-specific hOdf2-GFP transgenic mouse model, and relates to the technical field of reproductive biology and transgenic animal model construction. According to the construction method disclosed by the invention, based on a piggyBac transposon expression vector system, a DNA (Deoxyribose Nucleic Acid) sequence containing a coding human source Odf2 is effectively connected with a Protamine1 promoter. The transgenic positive mouse constructed by the method can specifically express human Odf2, has a dynamic change from sperm to zygote to eight cell stages, realizes early continuous visual tracking of embryos, has genetic stability, keeps expression and phenotype consistency across generations, and has a good application prospect. The method can be applied to the fields of environmental poison evaluation, asthenospermia mechanisms, drug screening and the like.

This invention discloses the application of the small molecule compound Z2114502286 in the preparation of drugs for the prevention and treatment of liver-related diseases. This invention reveals for the first time that Z2114502286 is a THBS2-specific small molecule inhibitor and verifies its feasibility in preventing and treating liver fibrosis by targeting and inhibiting the THBS2 protein. Through virtual screening and molecular docking, this compound can directly bind to the THBS2 protein. In vitro and in vivo studies show that this compound inhibits THBS2-mediated HSC activation in a concentration- and time-dependent manner, inhibits ECM deposition in a mouse model of liver fibrosis, downregulates the expression of fibrosis-related genes, and alleviates liver inflammation and macrophage infiltration. Simultaneously, the targeted efficacy was verified in a transgenic mouse model specifically expressing human THBS2, demonstrating significant clinical translational value. Furthermore, it exhibits bioavailability and safety at effective dosages, showing broad clinical application prospects.

The invention discloses a construction method of a Rosa26-APP-eta-CTF transgenic mouse model and an application of the Rosa26-APP-eta-CTF transgenic mouse model. According to the invention, a CRISPR / Cas9 mediated embryo gene editing technology is utilized, an expression cassette containing a human-derived APP-eta-CTF-mCherry fusion gene is inserted into a mouse Rosa26 site at a fixed point through a homologous recombination repair mechanism, and a transgenic mouse model capable of inducing expression is constructed. The model fills the blank of lack of an animal model for specifically simulating in-vivo accumulation of eta-CTF in the prior art, and provides a key tool for researching the effect of an eta-secretase pathway in the Alzheimer's disease.

The invention discloses a construction method of a microglial cell conditional knock-down dynactin animal model, which comprises the following steps: selecting a Dctn1LoxP gene knock-in mouse, hybridizing the Dctn1LoxP gene knock-in mouse with a Cx3cr1-Cre transgenic mouse, and carrying out three rounds of breeding to obtain a genotype Dctn1LoxP / LoxP; a mouse of Cx3cr1-Cre; the expression of Cre recombinase is driven by a Cx3cr1 gene promoter, and the 2nd to 4th exons of the Dctn1 gene are cut, so that the knockout of p150Glue and the reduction of the protein levels of other subunits of dynactin are realized. The model is strong in specificity and high in knock-down efficiency, animals show microglial cell morphology abnormity and movement coordination function defects, the model can be used for researching the influence of dynactin on microglial cell neurobiological functions, and a reliable tool is provided for pathogenesis exploration and drug screening of related neurological dysfunction.

The invention belongs to the field of bioengineering, and particularly relates to a construction method and application of a transgenic mouse for in-vivo screening of a T cell membrane protein gene. Aiming at the problems of low in-vivo screening research efficiency, library instability, tissue applicability limitation and the like of traditional T cell membrane protein genes, sgRNA library plasmids are constructed by designing sgRNA of a T cell membrane protein target gene and a control gene; three positive mice containing Cas9, Ubc-CreERT2 and sgRNA libraries are obtained through embryo microinjection and hybridization, genotype identification, inducer regulation and control, disease modeling, multi-tissue sample treatment and NGS analysis technical systems are established in a matched mode, and in-vivo multi-tissue sample high-throughput screening of T cell membrane protein genes is achieved. The model can guarantee library stability and screening reliability, covers multi-tissue research scenes, links in-vitro screening and in-vivo verification and is suitable for T cell membrane protein gene function analysis and disease target mining, the technical process can be popularized to other gene families, and systematic biological research is promoted.

The invention belongs to the field of biological medicine, and provides a construction method and application of an STK11 gene knockout male mouse model for simulating male breast cancer, and the construction method comprises the following steps: mixing an sgRNA composition with Cas9 protein to obtain a gene editing liquid; transferring the obtained gene editing liquid into a mouse fertilized egg, and then transplanting the fertilized egg into a pseudo-pregnant female mouse to obtain an F0-generation mouse; backcrossing the F0-generation mouse with the wild male mouse to obtain an F1-generation heterozygote mouse; the F1-generation heterozygote mice are intercrossed to obtain F2-generation homozygote male mice, the gene phenotype of the F2-generation homozygote male mice is STK11- / -, the F2-generation homozygote male mice are backcrossed with MMTV-PyMT transgenic mice to obtain the STK11 gene knockout male mouse model for simulating the male breast cancer, and the male mouse model can stably reproduce the typical pathological phenotype of occurrence and development of the male breast cancer and can be used for simulating the male breast cancer. The method can be applied to screening and / or evaluating candidate drugs for treating male breast cancer.

The invention discloses a construction method and application of a transgenic mouse model for in-vivo screening of immunotherapy target genes for regulating and controlling T cell metabolism, and belongs to the technical field of animal model construction. The technical problems to be solved are that in-vitro screening target spots for regulating and controlling T cell metabolism immunotherapy target genes in the prior art are poor in authenticity, complex in operation and high in cost, and corresponding in-vivo screening tools are lacked. According to the key points of the technical scheme, the construction method of the transgenic mouse model for in-vivo screening of the immunotherapy target genes for regulating and controlling T cell metabolism is provided, sgRNA (SEQ ID NO.1-150) of 140 immunotherapy target genes for regulating and controlling T cell metabolism and 10 control genes are designed, 150mer library plasmids are constructed step by step, and then model mice are constructed; target spots are screened through tamoxifen induction in combination with a tumor model, flow sorting and NGS analysis, 150 immunotherapy target genes for regulating and controlling T cell metabolism can be covered, operation is easy and efficient, and the screening cost is remarkably reduced.

The application belongs to the technical field of biology, and particularly relates to a transgenic animal model and a construction method and application thereof. flox / flox The transgenic mouse is mated with the Agc1-CreER transgenic mouse to generate the Axin1 Agc1ER Conditional knockout mouse, and then Axin1 Agc1ER The conditional knockout mouse Axin1 expression is down-regulated, and a disease combined model with pathological characteristics of growth plate chondrocyte hypertrophy, ectopic ossification and knee joint articular cartilage degeneration is successfully established; the Axin1 Agc1ER Conditional knockout mouse animal model has growth plate chondrocyte hypertrophy, ectopic ossification and osteoarthritis characteristics similar to the related pathological characteristics in clinic, and can be used for systematically researching the pathogenesis of growth plate chondrocyte hypertrophy, ectopic ossification and osteoarthritis and further screening drugs for treating related diseases.

The invention discloses a construction method and application of an animal model for expressing human ATP7B and a high-frequency mutant transgenic mouse of the human ATP7B, and belongs to the technical field of animal model construction. The invention provides a construction method of a human ATP7B high-frequency mutant transgenic mouse model. The method comprises the following steps: firstly, transforming a BAC plasmid, knocking out a non-target gene, and accurately introducing specific p.R778L, p.P992L and p.T935M point mutation of East Asian population by utilizing a CRISPR / Cas9 technology, so as to obtain mutant BAC DNA (Deoxyribose Nucleic Acid); and then, microinjection is carried out on the mouse fertilized eggs, and transgenic positive F0-generation mice are obtained through cultivation. And finally, hybridizing and screening with Atp7b gene knockout mice in multiple generations to obtain a target strain of which the endogenous Atp7b is completely deleted and which only expresses the human mutant ATP7B. The model can accurately simulate the genetic background and pathological phenotype of East Asia WD patients, and can be used for screening targeted drugs aiming at human ATP7B mutation and evaluating the improvement effect of the drugs on liver and nerve injury.

The invention discloses an anti-CHI3L2 neutralizing antibody and pharmaceutical application thereof in inflammatory diseases, and discovers that chitinase-like protein CHI3L2 is remarkably and highly expressed in plasma and tissues of atherosclerosis patients and is inducibly expressed in psoriasis skin lesion keratinocytes for the first time. The construction of a CHI3L2 transgenic mouse model proves that the CHI3L2 can promote the development of atherosclerotic plaques and skin inflammation response. The anti-CHI3L2 neutralizing antibody can inhibit THP-1 macrophage lipid uptake in vitro and inhibit HaCaT keratinocyte inflammatory factor secretion, and artery plaque load and IMQ-induced skin inflammation are remarkably relieved through intravenous or intradermal administration in vivo. Mechanism research shows that the antibody reduces release of factors such as IL-6, IL-8, TNF-alpha and the like by blocking a CHI3L2-mediated inflammation signal channel, and improves pathological changes of tissues. The neutralizing antibody provided by the invention can be used for preparing a pharmaceutical composition for treating atherosclerosis or psoriasis, has the treatment potential of'one target for double diseases', and has important clinical transformation value.

The invention discloses a construction method and application of a transgenic mouse model for in-vivo autoimmune gene screening, belongs to the technical field of animal model construction, and aims to solve the problems that in-vitro screening targets are not real, the repeatability is poor, the operation is complicated, the application range is narrow, and in-vivo screening tools for multiple autoimmune genes are lacked. The method comprises the following steps: designing sgRNA (SEQ ID NO: 1-150) of 140 autoimmune genes and 10 control genes, and constructing 150mer library plasmids step by step; after microinjection, the mouse copulates with Cas9 and UBC-CreERT2 mice to obtain a Sanyang mouse, tamoxifen induction is carried out, autoimmune related genes are screened by combining a tumor model, flow sorting and NGS analysis screening targets, 150 autoimmune related genes can be covered, the screening is real, the repeatability is high, the operation is simple and convenient, the application is wide, and a key tool is provided for autoimmune research and target development.

The invention discloses a construction method and application of a Clock gene site-directed mutagenesis mouse model, and belongs to the technical field of basic medical animal models. The construction method comprises the following steps: designing and constructing a gRNA vector, synthesizing a Donor Oligo sequence, and co-injecting the gRNA vector, a Cas9 vector and the synthesized Donor Oligo into a fertilized egg to obtain an F0-generation transgenic mouse; and mating the F0-generation mouse with the wild-type mouse to obtain an F1-generation transgenic mouse. Phenotype analysis on F1-generation transgenic mice finds that Clock gene site-directed mutagenesis mice show age-progressing heart failure phenotypes which are basically consistent with heart failure caused by aging in a natural physiological state, so that the mice can be used as a model for chronic heart failure diseases, and the pathogenic mechanism of the mice is expected to be deeply researched on the basis of the model; therefore, technical support is provided for prevention and treatment of the disease.

The application discloses a construction method and application of a Clock gene site-directed mutant mouse model, and belongs to the technical field of basic medical animal models. The construction method comprises the following steps: gRNA vector design and construction, synthesis of a Donor Oligo sequence, co-injection of the gRNA vector, a Cas9 vector and the synthesized Donor Oligo into a fertilized egg to obtain F0 generation transgenic mice; and crossbreeding of the F0 generation mice with wild type mice to obtain F1 generation transgenic mice. Phenotype analysis on the F1 generation transgenic mice shows that the Clock gene site-directed mutant mouse exhibits an age progression heart failure phenotype, which is basically consistent with heart failure caused by aging in a natural physiological state, and therefore, the mouse can be used as a model of chronic heart failure disease, and is expected to be used for in-depth research on the pathogenesis based on the model, thereby providing technical support for prevention and treatment of the disease.

The application provides a mouse fertilized ovum, wherein a sequence shown in SEQ ID NO. 1 in the fifth exon of a Trp53 gene is replaced by a fluorescent protein gene, and the individual developed from the fertilized ovum is a transgenic mouse for in-vivo labeling of p53 protein, and a construction method of the aforementioned fertilized ovum and transgenic mouse is provided. Through expression of the p53 and fluorescent label fusion protein, the application realizes accurate in-vivo labeling of the p53 protein, directly reflects the level of the p53 protein, and can track and analyze and sort abnormal cells by using flow cytometry according to the fluorescent protein label, so as to realize analysis and monitoring of tumor cell characteristics and tumor development process.