21 results about "Wild Type Mouse" patented technology

The invention discloses a construction method of a COL4A5-K229X point mutation X-linked Alport syndrome mouse model. The construction method comprises the following steps: aiming at c.685Agt of a No.12 exon of a mouse COL4A5 gene; carrying out T point mutation, and designing and preparing Cas9 mRNA, gRNA and a donor vector; the components are mixed and then microinjected into fertilized eggs of a C57BL / 6J mouse to obtain an F0-generation mouse; identifying the genotype through PCR (Polymerase Chain Reaction) amplification and Sanger sequencing, and screening positive mice; mating the positive F0-generation mice with the wild-type mice, and breeding F1-generation and subsequent generations; the phenotype of the model is further verified through qPCR, biochemical analysis, light microscopic examination, transmission electron microscope and immunofluorescence. The model constructed by the invention shows hematuria, proteinuria, azemia, podocyte loss and irregular thickening and layering of glomerular basement membrane, is consistent with phenotypes of human XLAS patients, and provides an animal model tool for analyzing pathogenesis and developing treatment strategies.

The application discloses application of TESC as an Alzheimer's disease prevention and treatment target and relates to the field of biological medicines. A TESC overexpression (TESC-OE) hippocampal neuron cell line is constructed through lentivirus transfection, and the result shows that TESC-OE has the ability to resist A beta-induced neuron apoptosis and synapse damage. A TESC overexpression model in the hippocampus of a mouse is constructed through stereotactic injection of an adeno-associated virus, and the result shows that the hippocampal gray matter volume in the brain of the TESC-OE mouse is significantly larger than that of a wild type mouse. A beta stereotactic modeling is performed on the basis of TESC overexpression in the hippocampus, and the result shows that the TESC-OE mouse can significantly resist A beta-induced hippocampal atrophy, learning and memory dysfunction and impaired synaptic plasticity. Immunoblotting analysis reveals the molecular mechanism that TESC can increase the expression of anti-apoptotic genes in the hippocampus and reduce the expression of pro-apoptotic genes to exert a neuroprotective effect.

The application provides a method for constructing a Bmp4 gene knock-in mouse model, and the method comprises the following steps: obtaining sgRNA12 shown in SEQ ID NO. 3; mixing the sgRNA12, Cas9 mRNA and a targeting vector shown in SEQ ID NO. 1, and then microinjecting into a mouse zygote to obtain F0 generation mice; selecting F0 generation positive mice in genotype identification results of the F0 generation mice, and mating the F0 generation positive mice with wild type mice to obtain F1 generation mice with stable genotypes; and screening gene targeting mice with correct recombination, and mating the gene targeting mice with retinal specific knock-out tool mice Six3-Cre to obtain Bmp4 gene knock-in model mice. The method has high gene editing efficiency, and the transgenic mouse can highly express human Bmp4 protein.

The invention discloses a construction method and application of a DNAJB6a gene specific knockout mouse model, and belongs to the technical field of gene engineering. Comprising the following steps: S1, designing gRNA sequences aiming at ninth to tenth exon regions of the mouse DNAJB6 gene; s2, in vitro transcription is carried out to prepare Cas9mRNA and gRNA; s3, carrying out microinjection on Cas9mRNA and gRNA (guide Ribonucleic Acid) into a fertilized egg of the mouse; s4, transplanting the fertilized ovum after injection into the body of a pseudo-pregnant female mouse to obtain an F0-generation mouse; s5, screening positive F0-generation mice through genotype identification, and mating the positive F0-generation mice with the wild type mice to obtain F1-generation heterozygote mice; s6, the F1-generation heterozygote mice are matched to obtain F2-generation homozygote mice, and the DNAJB6a gene specific knockout mouse model is established. The construction method for constructing the DNAJB6a gene knockout mouse model, provided by the invention, is simple and convenient to operate, short in construction process time and stable in genotype, and a DNAJB6a gene knockout mouse can be effectively obtained.

The application relates to a construction method of a conditional overexpression ZFP595 transgenic mouse model and application thereof, and relates to the technical field of transgenesis.The construction method comprises the following steps: constructing a homologous recombination vector containing a ZFP595 gene, wherein the vector comprises a 3.3 kb 5' homologous arm, CAG-LSL-Zfp595-HA-IRES-tdTomato-Wpre-pA and a 3.3 kb 3' homologous arm; microinjecting Cas9 mRNA, gRNA and the homologous recombination vector into a zygote of a recipient mouse to obtain F0 generation mice, and performing identification; mating the F0 generation mice identified as positive with wild type mice and backcrossing to obtain a mouse model for conditionally overexpressing ZFP595.The conditional overexpression ZFP595 mouse model constructed by the construction method can be crossed with various Cre mice to realize specific overexpression in different tissues, organs and cells.

The invention belongs to the technical field of biological medicines, and relates to application of Kir4.2 in treatment of renal fibrosis. The invention discovers that the expression level of the Kir4.2 protein (formed by KCNJ15 gene coding) in renal tissues of a fibrotic mouse is obviously lower than that of a wild type mouse for the first time, and the difference has statistical significance; moreover, after the Kir4.2 is specifically knocked out of the renal proximal tubule, renal fibrosis under a UUO and FA induction model can be aggravated, and the fibrosis process of human renal tubule epithelial cells induced by TGF-beta1 in vitro can be remarkably reversed by overexpression of the Kir4.2. Therefore, Kir4.2 can be used as a drug, a drug target or a target gene in gene therapy, is applied to prevention, alleviation and treatment of renal fibrosis, and can provide a new strategy for prevention and treatment of renal fibrosis. The invention further provides a construction method of the kidney proximal tubule specificity Kir4.2 knockout animal model, and the constructed animal model can be used for researching renal fibrosis pathogenesis and screening drugs for preventing, relieving or / and treating renal fibrosis and has a wide application prospect.

The application relates to the technical field of animal model construction, and particularly discloses a Sypl1 gene knockout colorectal cancer mouse model construction method and application, which comprises the following steps: synthesizing a specific target site gRNA of a Sypl1 gene, mixing Cas9 protein and the target site gRNA to obtain an injection compound, microinjecting the injection compound into mouse zygotes, and transplanting the surviving zygotes into the oviducts of pseudopregnant female mice to obtain F0 generation mice, and the mice born after 20 days are Sypl1 gene knockout mouse animal models; and a chemical induction modeling method is used to construct a colorectal cancer model in wild type mice and gene knockout mice. The Sypl1 gene knockout mouse is constructed by using a CRISPR / Cas mediated genome engineering technology Sypl1 combined with the most widely used colorectal cancer chemical inducer azoxymethane / dextran sodium sulfate, a new colorectal cancer mouse model is constructed, compared with a traditional chemical induction model, the tumor formation rate can be significantly improved, and the tumor formation time can be shortened, and a more efficient animal model is provided for colorectal cancer research.

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 invention relates to the technical field of animal model construction, and particularly discloses a construction method and application of a Sypl1 gene knockout colorectal cancer mouse model, and the construction method comprises the following steps: synthesizing a specific target site gRNA of a Sypl1 gene, uniformly mixing Cas9 protein and the target site gRNA to obtain an injection compound, microinjecting the injection compound into a mouse fertilized egg to obtain a Sypl1 gene knockout colorectal cancer mouse model. And transplanting the survival fertilized ova into the salpingtube of a pseudopregnant female mouse for 20 days to obtain a mouse which is an F0-generation mouse, obtaining a Sypl1 gene knockout mouse animal model, and constructing a colorectal cancer model in a wild type mouse and a gene knockout mouse by adopting a chemical induction modeling method. A Sypl1 gene knockout mouse is constructed through a CRISPR / Cas mediated genome engineering technology, a novel colorectal cancer mouse model is constructed in combination with a colorectal cancer chemical inducer azomethane / dextran sodium sulfate which is most widely applied at present, and compared with a traditional chemical induction model, the novel colorectal cancer mouse model has the advantages that the tumor formation rate can be remarkably increased, the tumor formation time can be shortened, and the tumor formation cost can be reduced. And a more efficient animal model is provided for colorectal cancer research.

The invention discloses an application of fetal calf serum exosome in preparation of a medicine for treating UV irradiation skin injury. When the high-activity exosome derived from the fetal calf serum is used for performing intervention treatment on a C57 wild type mouse UV irradiation skin injury model, the result shows that the exosome derived from the fetal calf serum can remarkably promote pathological improvement of the UV irradiation skin injury model and promote repair of ulceration caused by skin ultraviolet radiation; the regeneration of the epidermal layer and the corium layer of the skin is promoted. The effect of improving and treating the skin injury caused by UV irradiation is achieved, and no obvious side effect exists. Generally speaking, the obtained fetal calf serum-derived high-activity exosome can be used as an effective means for treating UV irradiation skin injury.

The invention discloses application of alpha1-acid glycoprotein in preparation of a medicine for treating lung injury caused by acute pancreatitis, and belongs to the technical field of biological medicine. An acute pancreatitis lung injury model is constructed through ORM1 gene knockout mice, the mice are divided into a normal group, a model group and a treatment group for research, target organ pathological injury degree detection, immunohistochemical analysis and serum inflammatory factor expression quantity are performed after injection administration, and the curative effect difference of the gene knockout mice and wild type mice is compared, so that the acute pancreatitis lung injury model is established. And the treatment effect of the alpha1-acid glycoprotein on the acute pancreatitis lung injury is verified. The invention provides a key basis for research, development and preparation of drugs for treating acute pancreatitis lung injury, and has important clinical application value.

The invention discloses a congenital cataract mouse model with RagA GTPase point mutation, a construction method and application, and belongs to the technical field of gene editing. The method comprises the following steps: aiming at a gRNA sequence of a mouse RagA GTPase gene, jointly introducing gRNA and Cas9 mRNA into a mouse fertilized egg, and transplanting the fertilized egg into the uterus of a pseudopregnant female mouse to obtain an F0-generation mouse carrying RagA GTPase point mutation; and hybridizing the F0 generation mouse with a wild type mouse, and screening to obtain an F1 generation homozygote mouse carrying RagA GTPase point mutation. According to the invention, a RagA GTPase Leu60Arg point mutation mouse model is constructed by using a CRISPR-Cas9 technology for the first time, and the RagA GTPase Leu60Arg point mutation mouse model can simulate clinical phenotypic characteristics of human congenital cataract and is used for pathogenesis research.

The application provides a premature aging mouse and application thereof. The application also provides a construction method of the premature aging mouse. Compared with a wild type mouse, the Lmna gene of the premature aging mouse has a mutation of A604V. The application also provides a nucleic acid molecule encoding Lmna, wherein the nucleic acid molecule corresponds to a mutation of C to T at position 1811 and a mutation of C to T at position 1827 in the nucleotide sequence shown in SEQ ID NO:1. The premature aging mouse constructed by the application has an aging phenotype at about 3 months after birth, and the life span can reach about 5-12 months, the window period for research is prolonged, and the practicability of the premature aging mouse is further increased.

The invention belongs to the technical field of biology, and particularly relates to a mouse model for systemic expression or tissue specific expression of KRAS-G12D-EGFP fusion protein and a preparation method thereof, and the preparation method comprises the following steps: a) screening gRNA target sites; b) constructing a homologous recombinant vector; c) mixing the homologous recombinant vector, gRNA and Cas9 mRNA, injecting the mixture into a mouse fertilized egg, and transplanting the mixture into a mouse blastocyst; d) carrying out PCR (Polymerase Chain Reaction) screening on the F0-generation mice to obtain correct homologous recombination positive mice; and e) mating the positive F0-generation mouse with a wild type mouse, breeding, and carrying out PCR screening to obtain an F1-generation recombinant positive mouse. A mouse model modified by the gene is stable in heredity and can be used for researching the biological function of the KRAS-G12D mutant protein in a mouse body. Particularly, the mouse model can be used for researching the function of the KRAS-G12D mutant protein at the tissue specificity level, and the mutant protein can be subjected to cell tissue tracing by using EGFP fluorescent protein, so that the mouse model has a certain application value.

The application discloses application of PDGF-C in corneal injury repair. The application research finds that in a corneal injury model mouse in which Pdgf-c is knocked out, Pdgf-c knockout reduces the area of corneal healing in the corneal injury mouse model, and the number of corneal limbal stem cells with proliferative activity is significantly less than that of wild type mice. Through AAV-mediated PDGF-C overexpression, the proliferation of corneal limbal stem cells can be significantly promoted, and the area of corneal healing in the corneal injury mouse model is significantly increased. These results show that overexpression of PDGF-C significantly improves the ability of corneal injury healing. Therefore, the application provides a new method for treating corneal injury.

The invention relates to the technical field of biotechnology, in particular to a two-leaf aortic valve disease model mouse as well as a construction method and application thereof, and the construction method comprises the following steps: designing gRNA of a targeted mouse FGFR3 exon 5-7, adapting a T7 promoter, and carrying out transcription purification after off-target prediction; mixing gRNA with Cas9, adding glycerol, incubating to obtain an RNP compound, and verifying the activity; rNP is injected into fertilized eggs, and after culture, the fertilized eggs are transplanted to pseudopregnant female rats to obtain an F0 generation; the F0 and a wild type mouse are subjected to cage combination to obtain F1, F1 heterozygotes are subjected to reproduction to obtain F2, and homozygotes are identified through PCR and sequencing; and taking F2 heart HE staining ornamental deformity, and detecting the expression of FGFR3 by RT-PCR and WB. According to the model, R-N type two-leaf aortic valve and right ventricle double-outlet deformity can be stably reproduced, the construction method is accurate and efficient, the model can be used for disease mechanism research and drug screening, and the blank of specific subtype research tools is filled.

The invention belongs to the technical field of biology, and discloses a construction method and application of a Pik3ip1 gene knockout mouse model. The construction method comprises the following steps: (1) transferring sgRNA1 (SEQ ID No.2), sgRNA2 (SEQ ID No.3) and Cas9 protein into a mouse fertilized egg, and carrying out development and delivery to obtain an F0-generation chimeric mouse; (2) hybridizing the F0-generation chimeric mouse with a wild type mouse to obtain an F1-generation heterozygote mouse; and (3) performing same-genotype male and female mating on the F1-generation heterozygote mice to obtain the Pik3ip1 gene knockout mouse model. According to the invention, the Pik3ip1 gene knockout mouse model is constructed by utilizing the combination of sgRNA and CRISPR / Cas9 technology, the construction method is stable, and the constructed Pik3ip1 gene knockout mouse model can be applied to animal models such as inflammation and is used for deeply researching the action and mechanism of the Pik3ip1 gene in diseases such as inflammation.

The invention provides a mouse model construction method for simulating the morbidity process of brain premature senility, ABAD knock-down is achieved in the hippocampus of a healthy adult mouse, and the pathological process of brain premature senility morbidity is simulated. According to the mouse model obtained by the invention, the expression quantity of the brain hippocampus ABAD gene is 30% or more lower than that of a wild type mouse, and the mouse model shows pathological characteristics which tend to be consistent with those of a mouse with brain premature senility.

The invention relates to a construction method and application of a conditional knockout mouse model for studying the influence of a CLCF1 gene on cortical bone development. GRNA, Cas9 mRNA and a targeting vector are microinjected into a fertilized egg of a C57BL / 6J mouse, the fertilized egg after injection is transplanted to a pseudo-pregnant female mouse, and an F0-generation mouse is produced; the method comprises the following steps: hybridizing a positive F0 generation mouse which is subjected to correct homologous recombination at a target site of the CLCF1 gene with a wild type mouse to obtain a flox heterozygote mouse of the CLCF1 gene; the CLCF1 gene flox heterozygote mice are subjected to mutual mating, and CLCF1 gene flox homozygote mice are obtained; a CLCF1 gene flox homozygote mouse and a Cd68-Cre tool mouse are subjected to mating, a mouse with the genotype of CLCF1flox / flox and carrying Cd68-Cre is obtained, and the mouse is the target CLCF1 conditional gene knockout mouse model. The CLCF1 conditional gene knockout mouse model disclosed by the invention is a powerful tool for researching functions of the CLCF1 gene. Meanwhile, the model also lays a solid foundation for simulating skeletal system diseases and screening prevention and treatment drugs for bone mass reduction, skeletal atrophy or dysplasia, and provides an important experimental platform.

The application relates to a method for constructing a Down syndrome mouse model capable of realizing conditional genetic correction and application, and comprises the following steps: inserting a LoxP sequence into the distal centromere end of the Zbtb21 gene at the end of the homologous region of human chromosome 21 on mouse chromosome 16 to obtain Zbtb21-LoxP / + F0 mice; crossing the Zbtb21-LoxP / + F0 mice with wild type mice to screen out Zbtb21-LoxP / + F1 mice; crossing the Zbtb21-LoxP / + mice with Dp16 / + mice to obtain Dp16 / Zbtb21-LoxP double-positive mice, and then crossing the Dp16 / Zbtb21-LoxP double-positive mice with wild type mice to obtain Dp16; Zbtb21-LoxP / + double-positive mice; and using a DS-Flox mouse model, combining with space-time expression regulation of Cre recombinase, the DS genetic defect can be corrected in specific target tissue cells or development stages.