CN110622921B - 阿尔茨海默病病变区FoxG1过表达小鼠模型的构建方法及应用 - Google Patents
阿尔茨海默病病变区FoxG1过表达小鼠模型的构建方法及应用 Download PDFInfo
- Publication number
- CN110622921B CN110622921B CN201910933563.9A CN201910933563A CN110622921B CN 110622921 B CN110622921 B CN 110622921B CN 201910933563 A CN201910933563 A CN 201910933563A CN 110622921 B CN110622921 B CN 110622921B
- Authority
- CN
- China
- Prior art keywords
- foxg1
- mouse
- overexpression
- alzheimer disease
- alzheimer
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Active
Links
- 101710087964 Forkhead box protein G1 Proteins 0.000 title claims abstract description 127
- 208000024827 Alzheimer disease Diseases 0.000 title claims abstract description 95
- 230000002018 overexpression Effects 0.000 title claims abstract description 75
- 230000003902 lesion Effects 0.000 title claims abstract description 50
- 238000010172 mouse model Methods 0.000 title claims abstract description 35
- 238000010276 construction Methods 0.000 title claims abstract description 16
- 241000699666 Mus <mouse, genus> Species 0.000 claims abstract description 74
- 210000004556 brain Anatomy 0.000 claims abstract description 68
- 241000699670 Mus sp. Species 0.000 claims abstract description 46
- NKANXQFJJICGDU-QPLCGJKRSA-N Tamoxifen Chemical compound C=1C=CC=CC=1C(/CC)=C(C=1C=CC(OCCN(C)C)=CC=1)/C1=CC=CC=C1 NKANXQFJJICGDU-QPLCGJKRSA-N 0.000 claims abstract description 26
- 210000001577 neostriatum Anatomy 0.000 claims abstract description 24
- 210000001320 hippocampus Anatomy 0.000 claims abstract description 22
- 238000000034 method Methods 0.000 claims abstract description 22
- 229940079593 drug Drugs 0.000 claims abstract description 13
- 239000003814 drug Substances 0.000 claims abstract description 13
- 229960001603 tamoxifen Drugs 0.000 claims abstract description 13
- 230000001054 cortical effect Effects 0.000 claims abstract description 9
- 230000000971 hippocampal effect Effects 0.000 claims abstract description 8
- 230000013011 mating Effects 0.000 claims abstract description 8
- 108010051219 Cre recombinase Proteins 0.000 claims abstract description 6
- 238000002360 preparation method Methods 0.000 claims abstract description 4
- 108090000623 proteins and genes Proteins 0.000 claims description 15
- 238000011830 transgenic mouse model Methods 0.000 claims description 13
- 238000009395 breeding Methods 0.000 claims description 11
- 230000001488 breeding effect Effects 0.000 claims description 11
- 230000014509 gene expression Effects 0.000 claims description 9
- 239000002285 corn oil Substances 0.000 claims description 8
- 235000005687 corn oil Nutrition 0.000 claims description 8
- 241000283707 Capra Species 0.000 claims description 7
- 238000012216 screening Methods 0.000 claims description 5
- 101000892837 Mus musculus Forkhead box protein G1 Proteins 0.000 claims description 4
- 230000001939 inductive effect Effects 0.000 claims description 4
- 102100027499 5-hydroxytryptamine receptor 1B Human genes 0.000 claims description 3
- 101710138639 5-hydroxytryptamine receptor 1B Proteins 0.000 claims description 3
- 238000010166 immunofluorescence Methods 0.000 claims description 3
- 230000001575 pathological effect Effects 0.000 claims description 3
- 241000283973 Oryctolagus cuniculus Species 0.000 claims description 2
- 230000008859 change Effects 0.000 claims description 2
- 230000036285 pathological change Effects 0.000 claims 2
- 230000037396 body weight Effects 0.000 claims 1
- 230000007246 mechanism Effects 0.000 abstract description 5
- 238000011161 development Methods 0.000 abstract description 4
- 238000011160 research Methods 0.000 abstract description 4
- 230000009261 transgenic effect Effects 0.000 abstract 1
- 108020004414 DNA Proteins 0.000 description 28
- 238000003752 polymerase chain reaction Methods 0.000 description 25
- 101150036876 cre gene Proteins 0.000 description 22
- 102000013455 Amyloid beta-Peptides Human genes 0.000 description 8
- 108010090849 Amyloid beta-Peptides Proteins 0.000 description 8
- 210000004027 cell Anatomy 0.000 description 7
- 238000010586 diagram Methods 0.000 description 7
- 210000003371 toe Anatomy 0.000 description 7
- 239000000243 solution Substances 0.000 description 5
- 238000011144 upstream manufacturing Methods 0.000 description 5
- 241000699660 Mus musculus Species 0.000 description 4
- 210000005013 brain tissue Anatomy 0.000 description 4
- 238000005516 engineering process Methods 0.000 description 4
- 239000003147 molecular marker Substances 0.000 description 4
- 210000002442 prefrontal cortex Anatomy 0.000 description 4
- 238000005406 washing Methods 0.000 description 4
- PEDCQBHIVMGVHV-UHFFFAOYSA-N Glycerine Chemical compound OCC(O)CO PEDCQBHIVMGVHV-UHFFFAOYSA-N 0.000 description 3
- 238000012408 PCR amplification Methods 0.000 description 3
- SXEHKFHPFVVDIR-UHFFFAOYSA-N [4-(4-hydrazinylphenyl)phenyl]hydrazine Chemical compound C1=CC(NN)=CC=C1C1=CC=C(NN)C=C1 SXEHKFHPFVVDIR-UHFFFAOYSA-N 0.000 description 3
- 210000000683 abdominal cavity Anatomy 0.000 description 3
- 230000005856 abnormality Effects 0.000 description 3
- 230000018109 developmental process Effects 0.000 description 3
- 230000001965 increasing effect Effects 0.000 description 3
- 239000000463 material Substances 0.000 description 3
- 230000004048 modification Effects 0.000 description 3
- 238000012986 modification Methods 0.000 description 3
- 230000003797 telogen phase Effects 0.000 description 3
- 238000013518 transcription Methods 0.000 description 3
- 230000035897 transcription Effects 0.000 description 3
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 description 2
- WZUVPPKBWHMQCE-UHFFFAOYSA-N Haematoxylin Chemical compound C12=CC(O)=C(O)C=C2CC2(O)C1C1=CC=C(O)C(O)=C1OC2 WZUVPPKBWHMQCE-UHFFFAOYSA-N 0.000 description 2
- MHAJPDPJQMAIIY-UHFFFAOYSA-N Hydrogen peroxide Chemical compound OO MHAJPDPJQMAIIY-UHFFFAOYSA-N 0.000 description 2
- 241001465754 Metazoa Species 0.000 description 2
- 241001529936 Murinae Species 0.000 description 2
- 229930040373 Paraformaldehyde Natural products 0.000 description 2
- 108091023040 Transcription factor Proteins 0.000 description 2
- 102000040945 Transcription factor Human genes 0.000 description 2
- 238000010171 animal model Methods 0.000 description 2
- 239000000427 antigen Substances 0.000 description 2
- 102000036639 antigens Human genes 0.000 description 2
- 108091007433 antigens Proteins 0.000 description 2
- 230000033228 biological regulation Effects 0.000 description 2
- 230000015572 biosynthetic process Effects 0.000 description 2
- 238000006243 chemical reaction Methods 0.000 description 2
- 238000001816 cooling Methods 0.000 description 2
- 238000001962 electrophoresis Methods 0.000 description 2
- 238000001502 gel electrophoresis Methods 0.000 description 2
- 239000008187 granular material Substances 0.000 description 2
- 238000011532 immunohistochemical staining Methods 0.000 description 2
- 238000003364 immunohistochemistry Methods 0.000 description 2
- 230000000415 inactivating effect Effects 0.000 description 2
- 238000002347 injection Methods 0.000 description 2
- 239000007924 injection Substances 0.000 description 2
- 230000004770 neurodegeneration Effects 0.000 description 2
- 229920002866 paraformaldehyde Polymers 0.000 description 2
- 239000000047 product Substances 0.000 description 2
- 230000002062 proliferating effect Effects 0.000 description 2
- 102000004169 proteins and genes Human genes 0.000 description 2
- QKNYBSVHEMOAJP-UHFFFAOYSA-N 2-amino-2-(hydroxymethyl)propane-1,3-diol;hydron;chloride Chemical compound Cl.OCC(N)(CO)CO QKNYBSVHEMOAJP-UHFFFAOYSA-N 0.000 description 1
- FWBHETKCLVMNFS-UHFFFAOYSA-N 4',6-Diamino-2-phenylindol Chemical compound C1=CC(C(=N)N)=CC=C1C1=CC2=CC=C(C(N)=N)C=C2N1 FWBHETKCLVMNFS-UHFFFAOYSA-N 0.000 description 1
- FWMNVWWHGCHHJJ-SKKKGAJSSA-N 4-amino-1-[(2r)-6-amino-2-[[(2r)-2-[[(2r)-2-[[(2r)-2-amino-3-phenylpropanoyl]amino]-3-phenylpropanoyl]amino]-4-methylpentanoyl]amino]hexanoyl]piperidine-4-carboxylic acid Chemical compound C([C@H](C(=O)N[C@H](CC(C)C)C(=O)N[C@H](CCCCN)C(=O)N1CCC(N)(CC1)C(O)=O)NC(=O)[C@H](N)CC=1C=CC=CC=1)C1=CC=CC=C1 FWMNVWWHGCHHJJ-SKKKGAJSSA-N 0.000 description 1
- 101710137189 Amyloid-beta A4 protein Proteins 0.000 description 1
- 101710151993 Amyloid-beta precursor protein Proteins 0.000 description 1
- 101100298998 Caenorhabditis elegans pbs-3 gene Proteins 0.000 description 1
- 238000007400 DNA extraction Methods 0.000 description 1
- 108010067770 Endopeptidase K Proteins 0.000 description 1
- 208000013548 FOXG1 disease Diseases 0.000 description 1
- 101150026630 FOXG1 gene Proteins 0.000 description 1
- 108090000852 Forkhead Transcription Factors Proteins 0.000 description 1
- 102000004315 Forkhead Transcription Factors Human genes 0.000 description 1
- 208000029726 Neurodevelopmental disease Diseases 0.000 description 1
- CTQNGGLPUBDAKN-UHFFFAOYSA-N O-Xylene Chemical compound CC1=CC=CC=C1C CTQNGGLPUBDAKN-UHFFFAOYSA-N 0.000 description 1
- 108091005804 Peptidases Proteins 0.000 description 1
- 102000035195 Peptidases Human genes 0.000 description 1
- 102000003992 Peroxidases Human genes 0.000 description 1
- 206010034719 Personality change Diseases 0.000 description 1
- 229920001213 Polysorbate 20 Polymers 0.000 description 1
- 108010091086 Recombinases Proteins 0.000 description 1
- 102000018120 Recombinases Human genes 0.000 description 1
- 208000006289 Rett Syndrome Diseases 0.000 description 1
- 229930006000 Sucrose Natural products 0.000 description 1
- CZMRCDWAGMRECN-UGDNZRGBSA-N Sucrose Chemical compound O[C@H]1[C@H](O)[C@@H](CO)O[C@@]1(CO)O[C@@H]1[C@H](O)[C@@H](O)[C@H](O)[C@@H](CO)O1 CZMRCDWAGMRECN-UGDNZRGBSA-N 0.000 description 1
- 239000011543 agarose gel Substances 0.000 description 1
- 238000000246 agarose gel electrophoresis Methods 0.000 description 1
- 230000032683 aging Effects 0.000 description 1
- 230000003321 amplification Effects 0.000 description 1
- 230000003376 axonal effect Effects 0.000 description 1
- 230000009286 beneficial effect Effects 0.000 description 1
- 230000000903 blocking effect Effects 0.000 description 1
- 210000004438 cajal-retzius cell Anatomy 0.000 description 1
- 230000005754 cellular signaling Effects 0.000 description 1
- 210000003169 central nervous system Anatomy 0.000 description 1
- PBAYDYUZOSNJGU-UHFFFAOYSA-N chelidonic acid Natural products OC(=O)C1=CC(=O)C=C(C(O)=O)O1 PBAYDYUZOSNJGU-UHFFFAOYSA-N 0.000 description 1
- 239000003153 chemical reaction reagent Substances 0.000 description 1
- 208000010877 cognitive disease Diseases 0.000 description 1
- 230000007547 defect Effects 0.000 description 1
- 230000003412 degenerative effect Effects 0.000 description 1
- 230000018044 dehydration Effects 0.000 description 1
- 238000006297 dehydration reaction Methods 0.000 description 1
- 210000001947 dentate gyrus Anatomy 0.000 description 1
- 230000008021 deposition Effects 0.000 description 1
- 238000001514 detection method Methods 0.000 description 1
- 208000037265 diseases, disorders, signs and symptoms Diseases 0.000 description 1
- 238000009509 drug development Methods 0.000 description 1
- 210000003027 ear inner Anatomy 0.000 description 1
- 206010015037 epilepsy Diseases 0.000 description 1
- 230000020339 forebrain development Effects 0.000 description 1
- 210000003194 forelimb Anatomy 0.000 description 1
- 238000007710 freezing Methods 0.000 description 1
- 230000008014 freezing Effects 0.000 description 1
- 230000006870 function Effects 0.000 description 1
- 230000004927 fusion Effects 0.000 description 1
- 230000012010 growth Effects 0.000 description 1
- 230000009808 hippocampal neurogenesis Effects 0.000 description 1
- 230000006801 homologous recombination Effects 0.000 description 1
- 238000002744 homologous recombination Methods 0.000 description 1
- 238000010191 image analysis Methods 0.000 description 1
- 238000012151 immunohistochemical method Methods 0.000 description 1
- 230000006698 induction Effects 0.000 description 1
- 239000003112 inhibitor Substances 0.000 description 1
- 238000002955 isolation Methods 0.000 description 1
- 210000003141 lower extremity Anatomy 0.000 description 1
- 239000012139 lysis buffer Substances 0.000 description 1
- 230000001404 mediated effect Effects 0.000 description 1
- 108020004999 messenger RNA Proteins 0.000 description 1
- 208000004141 microcephaly Diseases 0.000 description 1
- 230000035772 mutation Effects 0.000 description 1
- 210000005036 nerve Anatomy 0.000 description 1
- 208000015122 neurodegenerative disease Diseases 0.000 description 1
- 230000007472 neurodevelopment Effects 0.000 description 1
- 210000002682 neurofibrillary tangle Anatomy 0.000 description 1
- 210000002569 neuron Anatomy 0.000 description 1
- 230000007996 neuronal plasticity Effects 0.000 description 1
- 238000003199 nucleic acid amplification method Methods 0.000 description 1
- 239000012188 paraffin wax Substances 0.000 description 1
- 230000007170 pathology Effects 0.000 description 1
- 108040007629 peroxidase activity proteins Proteins 0.000 description 1
- 239000000256 polyoxyethylene sorbitan monolaurate Substances 0.000 description 1
- 235000010486 polyoxyethylene sorbitan monolaurate Nutrition 0.000 description 1
- 230000000750 progressive effect Effects 0.000 description 1
- 230000000644 propagated effect Effects 0.000 description 1
- 235000019833 protease Nutrition 0.000 description 1
- 238000005215 recombination Methods 0.000 description 1
- 230000022532 regulation of transcription, DNA-dependent Effects 0.000 description 1
- 230000001105 regulatory effect Effects 0.000 description 1
- 230000002207 retinal effect Effects 0.000 description 1
- 238000007789 sealing Methods 0.000 description 1
- 230000009758 senescence Effects 0.000 description 1
- 230000007507 senile plaque formation Effects 0.000 description 1
- 208000022288 senile plaque formation Diseases 0.000 description 1
- 210000002966 serum Anatomy 0.000 description 1
- 239000002689 soil Substances 0.000 description 1
- 239000005720 sucrose Substances 0.000 description 1
- 239000006228 supernatant Substances 0.000 description 1
- 210000001519 tissue Anatomy 0.000 description 1
- 238000012800 visualization Methods 0.000 description 1
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 description 1
- 239000001993 wax Substances 0.000 description 1
- 239000008096 xylene Substances 0.000 description 1
Images
Classifications
-
- A—HUMAN NECESSITIES
- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
- A01K—ANIMAL HUSBANDRY; AVICULTURE; APICULTURE; PISCICULTURE; FISHING; REARING OR BREEDING ANIMALS, NOT OTHERWISE PROVIDED FOR; NEW BREEDS OF ANIMALS
- A01K67/00—Rearing or breeding animals, not otherwise provided for; New or modified breeds of animals
- A01K67/027—New or modified breeds of vertebrates
- A01K67/0275—Genetically modified vertebrates, e.g. transgenic
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K49/00—Preparations for testing in vivo
- A61K49/0004—Screening or testing of compounds for diagnosis of disorders, assessment of conditions, e.g. renal clearance, gastric emptying, testing for diabetes, allergy, rheuma, pancreas functions
- A61K49/0008—Screening agents using (non-human) animal models or transgenic animal models or chimeric hosts, e.g. Alzheimer disease animal model, transgenic model for heart failure
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N15/00—Mutation or genetic engineering; DNA or RNA concerning genetic engineering, vectors, e.g. plasmids, or their isolation, preparation or purification; Use of hosts therefor
- C12N15/09—Recombinant DNA-technology
- C12N15/87—Introduction of foreign genetic material using processes not otherwise provided for, e.g. co-transformation
- C12N15/90—Stable introduction of foreign DNA into chromosome
-
- A—HUMAN NECESSITIES
- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
- A01K—ANIMAL HUSBANDRY; AVICULTURE; APICULTURE; PISCICULTURE; FISHING; REARING OR BREEDING ANIMALS, NOT OTHERWISE PROVIDED FOR; NEW BREEDS OF ANIMALS
- A01K2227/00—Animals characterised by species
- A01K2227/10—Mammal
- A01K2227/105—Murine
-
- A—HUMAN NECESSITIES
- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
- A01K—ANIMAL HUSBANDRY; AVICULTURE; APICULTURE; PISCICULTURE; FISHING; REARING OR BREEDING ANIMALS, NOT OTHERWISE PROVIDED FOR; NEW BREEDS OF ANIMALS
- A01K2267/00—Animals characterised by purpose
- A01K2267/03—Animal model, e.g. for test or diseases
- A01K2267/0306—Animal model for genetic diseases
- A01K2267/0312—Animal model for Alzheimer's disease
Landscapes
- Life Sciences & Earth Sciences (AREA)
- Health & Medical Sciences (AREA)
- Genetics & Genomics (AREA)
- Engineering & Computer Science (AREA)
- Zoology (AREA)
- Biotechnology (AREA)
- General Health & Medical Sciences (AREA)
- Biomedical Technology (AREA)
- Organic Chemistry (AREA)
- Chemical & Material Sciences (AREA)
- Environmental Sciences (AREA)
- General Engineering & Computer Science (AREA)
- Molecular Biology (AREA)
- Wood Science & Technology (AREA)
- Bioinformatics & Cheminformatics (AREA)
- Animal Behavior & Ethology (AREA)
- Veterinary Medicine (AREA)
- Plant Pathology (AREA)
- Endocrinology (AREA)
- Physics & Mathematics (AREA)
- Microbiology (AREA)
- Mycology (AREA)
- Biodiversity & Conservation Biology (AREA)
- Biochemistry (AREA)
- Animal Husbandry (AREA)
- Diabetes (AREA)
- Biophysics (AREA)
- Gastroenterology & Hepatology (AREA)
- Pathology (AREA)
- Rheumatology (AREA)
- Toxicology (AREA)
- Urology & Nephrology (AREA)
- Epidemiology (AREA)
- Public Health (AREA)
- Measuring Or Testing Involving Enzymes Or Micro-Organisms (AREA)
- Medicines That Contain Protein Lipid Enzymes And Other Medicines (AREA)
Abstract
本发明属于生物技术领域,具体涉及阿尔茨海默病病变区FoxG1过表达小鼠模型的构建方法及应用。本发明将FoxG1转基因小鼠与海马、皮层及纹状体特异脑区表达Cre重组酶的Cre工具小鼠交配,获得海马、皮层及纹状体FoxG1条件性过表达小鼠,再与阿尔茨海默病模型小鼠交配得到阿尔茨海默病病变脑区FoxG1条件性过表达小鼠。他莫昔芬注射诱其海马、皮层及纹状体病变脑区FoxG1蛋白过表达后制备在阿尔茨海默病模型鼠病变脑区FoxG1过表达小鼠。实现特定脑区FoxG1条件性过表达小鼠模型的制备,对阿尔茨海默病的机制研究及靶向药物开发具有重要的意义。
Description
技术领域
本发明属于生物技术领域,具体涉及阿尔茨海默病病变区FoxG1过表达小鼠模型的构建方法及应用。
背景技术
阿尔茨海默病(Alzheimer's disease,AD)作为一种中枢神经系统退行性病变,常发生于老年前期或老年期,主要以进行性的记忆认知功能障碍、人格性格改变为特征,其主要病理特为老年斑形成、神经原纤维缠结沉积、海马椎体细胞颗粒空泡变性以及神经元缺失等。随着社会老龄化的发展,AD的发病率逐年升高,当前已经成为严重的社会问题。然而,迄今为止,AD的发病原因及其发病机制尚不明确。越来越多的证据表明转录调控异常与AD发生密切相关,无论在AD细胞模型、转基因小鼠模型还是AD患者脑组织研究结果均证实mRNA转录水平的异常。
叉头框(Fox)转录因子家族是一类转录因子蛋白家族。其中,FoxG1是Fox家族的一个重要成员,执行着转录抑制功能。有研究表明,FoxG1突变与许多神经发育障碍如Rett综合征、癫痫及小头症密切相关。FoxG1作为端脑特异性的转录抑制因子,在胚胎期调控小鼠端脑的形成,在脑发育早期,FoxG1主要选择性表达于端脑形成的增殖细胞群,FoxG1使细胞持续维持在增殖状态,并阻止其分化为神经元,参与神经可塑性的调控;出生后FoxG1继续表达于海马齿状回和室管膜下区,调节出生后海马的神经形成。FoxG1促进视网膜轴突的生长,调控前脑的发育以及内耳和嗅觉系统的形成。然而FoxG1与AD的关系却鲜见报道,阿尔茨海默病病变脑区FoxG1过表达小鼠模型的制备还处于空缺状态。
发明内容
本发明针对现有技术中存在的不足,提供一种阿尔茨海默病病变区FoxG1过表达小鼠模型的构建方法及应用。
本发明通过基因重组技术构建阿尔茨海默病病变脑区FoxG1过表达小鼠,并对阿尔茨海默病病变脑区FoxG1过表达小鼠的鉴定及繁殖进行探讨,通过PCR、免疫组化染色(Immunohischemistry,IHC)及免疫荧光技术(Immunofluorescence,IF)验证阿尔茨海默病病变脑区FoxG1过表达小鼠模型的建立及FoxG1的蛋白表达,为与FoxG1相关的阿尔茨海默病的机制研究提供一个可信的动物模型。
本发明提供一种阿尔茨海默病病变脑区FoxG1过表达小鼠模型。
本发明还提供一种阿尔茨海默病病变脑区FoxG1过表达小鼠模型的构建方法。
为达到上述目的,本发明采取如下技术手段:
本发明将5-HT1B作为海马、皮层及纹状体特定脑区的启动子,利用Cre/Loxp重组酶系统及他莫昔芬诱导成功构建了阿尔茨海默病病变脑区FoxG1过表达小鼠,通过5-HT1B启动子驱动Cre重组酶在海马、皮层及纹状体特定脑区特异性表达,选择性敲除FoxG1转基因小鼠CAG-loxp-stop-loxp-FoxG1-IRES-EGFP中两个loxp基因中间的stop位点,诱导App-ps1小鼠海马、皮层及纹状体FoxG1条件性过表达,进而发现构建的阿尔茨海默病病变脑区FoxG1过表达小鼠中FoxG1存在高表达性。
本发明所述的阿尔茨海默病病变脑区FoxG1过表达小鼠模型的构建方法,具体包括如下步骤:
(1)FoxG1转基因小鼠与Cre工具小鼠交配繁育后获得海马、皮层及纹状体FoxG1条件性过表达小鼠;
(2)将繁育获得的海马、皮层及纹状体FoxG1条件性过表达小鼠与阿尔茨海默病模型小鼠,迅速扩大种群,得到阿尔茨海默病病变脑区FoxG1过表达小鼠;
(3)他莫昔芬/玉米油溶液诱导阿尔茨海默病病变脑区FoxG1过表达小鼠过表达FoxG1蛋白。
本发明还提供了一种对构建的阿尔茨海默病病变脑区FoxG1过表达小鼠进行鉴定的方法,利用特异性引物通过PCR方法进行多重PCR鉴定,所述海马、皮层及纹状体FoxG1条件性过表达小鼠的PCR引物序列如SEQ ID NO:5~10所示,所述阿尔茨海默病病变脑区FoxG1过表达小鼠的PCR引物序列如SEQ ID NO:1~10所示。
饲养获得的阿尔茨海默病病变脑区FoxG1过表达小鼠,生长至6个月,腹腔注射他莫昔芬/玉米油溶液,隔天注射一次,连续两周,诱导阿尔茨海默病病变脑区FoxG1过表达小鼠的海马、皮层及纹状体病变脑区过表达FoxG1蛋白。利用免疫荧光方法对阿尔茨海默病病变脑区FoxG1过表达小鼠进行FoxG1蛋白过表达鉴定。
优选的,FoxG1蛋白过表达鉴定中一抗为兔源GFP,二抗为山羊抗兔IgG。
本发明还提供了一种构建的阿尔茨海默病病变脑区FoxG1过表达小鼠模型在制备或筛选阿尔茨海默病治疗药物中的应用。
本发明还提供一种筛选阿尔茨海默病治疗药物的方法,所述方法为将阿尔茨海默病治疗备选药物施用于上述构建方法获得的阿尔茨海默病病变脑区FoxG1过表达小鼠模型的病状部位,通过比较用药前后小鼠病变脑区FoxG1过表达量的变化,筛选出使病变脑区FoxG1过表达量减少的药物作为阿尔茨海默病治疗药物。
与现有技术相比,本发明的有益效果是:
本发明将FoxG1转基因小鼠与在海马、皮层和纹状体特定脑区表达Cre重组酶的Cre工具小鼠杂交后获得海马、皮层及纹状体FoxG1条件性过表达小鼠,进一步与阿尔茨海默病模型小鼠交配后扩大种群,通过他莫昔芬注射诱导Cre介导的同源重组,移除两个loxp基因中的stop位点,使FoxG1选择性过表达于阿尔茨海默病的病变脑区,制备得到在阿尔茨海默病病变脑区FoxG1过表达的小鼠模型。实现了海马、皮层及纹状体在特定脑区FoxG1条件性过表达小鼠模型的构建,对FoxG1相关的阿尔茨海默病的机制研究及靶向药物开发具有重要的意义,并为神经发育及神经退行性疾病的机制研究提供一个可信的动物模型。
附图说明
图1是阿尔茨海默病病变脑区FoxG1过表达小鼠的构建方案示意图;
图2是阿尔茨海默病模型小鼠的PCR基因鉴定结果图;编号1~7代表阿尔茨海默病模型小鼠个体;
图3是阿尔茨海默病模型小鼠App-ps1的β-淀粉样蛋白(β-Amyloid)的免疫组化结果图;图中,(a)为2个月的App-ps1小鼠,(b)为6个月的App-ps1小鼠;
图4是繁育获得的海马、皮层及纹状体FoxG1条件性过表达小鼠的PCR基因鉴定结果图,M为DNA分子标记Marker;编号1~8代表海马、皮层及纹状体FoxG1条件性过表达小鼠个体;图中,(a)是FoxG1基因的目的条带,(b)是Cre基因的目的条带;
图5是繁育获得的阿尔茨海默病病变脑区FoxG1过表达小鼠的三重PCR基因鉴定结果图,M为DNA分子标记Marker;编号1~15代表阿尔茨海默病病变脑区FoxG1条件性过表达小鼠个体;图中(a)是Ps1和App的目的条带,(b)是FoxG1的目的条带,(c)是Cre的目的条带;
图6是野生型小鼠与阿尔茨海默病病变脑区FoxG1过表达小鼠前额皮层FoxG1表达GFP荧光鉴定结果对比图;图中,A是野生型小鼠,B是阿尔茨海默病病变脑区FoxG1过表达小鼠。
具体实施方式
本发明公开了一种阿尔茨海默病病变区FoxG1过表达小鼠模型的构建方法及应用,本领域技术人员可以借鉴本文内容,适当改进工艺参数实现。特别需要指出的是,所有类似的替换和改动对本领域技术人员来说是显而易见的,它们都被视为包括在本发明。本发明的方法及应用已经通过较佳实施例进行了描述,相关人员明显能在不脱离本发明内容、精神和范围内对本文所述的方法和应用进行改动或适当变更与组合,来实现和应用本发明技术。以下实施案例中的方法、设备、材料,如果未进行特别说明,均为本领域常规方法、设备和材料,均可由市场购得。
材料及试剂:
FoxG1转基因小鼠(CAG-loxp-stop-loxp-FoxG1-IRES-EGFP),品系名称B6/FVB,由东南大学赵春杰教授惠赠(Forced expression of Foxg1 in the Cortical Hem Leadsto the Transformation of Cajal-Retzius Cells into Dentate Granule Neurons),雌雄各2只;
Cre工具小鼠(B6; 129-5-HT1Btm1(CreERT)/Nju),从南京模式动物研究中心引进[许可证编号:SCxk(苏)2018-0008],雌雄各2只;
阿尔茨海默病模型小鼠(App-ps1),由南京模式动物研究中心引进,品系名称B6C3-Tg(APPswePSEN1dE9)/Nju,雄性2只,雌性6只;
玉米油(A0395699,ACROS),他莫昔芬(# T5648,Sigma),GFP抗体Bioss(兔-GFPcatalog # bs-0844R)、山羊抗兔IgG二抗(碧云天,catalog # p0186-1),鼠源β-Amyloid(Cell Signaling Technology, catalog # 15126),二抗山羊抗鼠IgG二抗(澳泉医疗科技有限公司,catalog # RQ7025);
转基因小鼠的饲养按照SPF动物饲养标准执行,经隔离观察未见异常后进入饲养区,严格按照SPF动物管理相关规定进行实验操作。
实施例一:阿尔茨海默病病变脑区FoxG1过表达小鼠模型的构建
参考附图1所示构建阿尔茨海默病病变脑区FoxG1过表达小鼠模型,具体构建方法包括如下步骤:
(1)Cre工具小鼠B6;129-5-HT1Btm1 (CreERT) /Nju脑部的海马、皮层和纹状体特定脑区可表达Cre重组酶,取FoxG1转基因小鼠CAG-loxp-stop-loxp-FoxG1-IRES-EGFP与Cre工具小鼠B6;129-5-HT1Btm1 (CreERT) /Nju交配,获得海马、皮层及纹状体FoxG1条件性过表达小鼠5-HT1Btm1(CreERT)/Nju ; FoxG1;
(2)将繁育获得的海马、皮层及纹状体FoxG1条件性过表达小鼠与阿尔茨海默病模型小鼠App-ps1交配,迅速扩大种群,得到阿尔茨海默病病变脑区FoxG1条件性过表达小鼠App-/Ps1;5-HT1Btm1(CreERT)/Nju; FoxG1;
(3)阿尔茨海默病病变脑区FoxG1过表达小鼠生长至6个月,腹腔注射他莫昔芬/玉米油溶液(75 mg/kg),隔天注射一次,连续两周,诱导小鼠海马、皮层及纹状体病变脑区过表达FoxG1蛋白。
实施例二:转基因小鼠的鉴定
采取剪去脚趾编号方法(剪取出生后5-6天的子鼠的脚趾),从后肢到前肢,从右到左依次编号,剪去的脚趾组织用于提取基因组DNA,基因组DNA提取大致步骤如下:
(1)取小鼠脚趾(按1-10的顺序)放入1.5 mL EP管中,适当离心30 s使其沉于管底;
(2)加入30 μl裂解液(0.5% 20% Tween-20, 1M KCL 50 mM,1M MgCl215 mM, 1MTris-HCl 2.5 mM, PH8.0,用前加200 μg/mL蛋白酶K 3 μL);
(3)55 ℃消化3-5 h, 12,000 rpm离心2 min, 95 ℃,15 min,灭活蛋白酶K,12,000 rpm离心2 min,所获得的上清中DNA用于PCR模板,-20 ℃保存,用于基因型鉴定。
、阿尔茨海默病模型小鼠App-ps1的鉴定
阿尔茨海默病模型小鼠App-ps1基因型中含有衰老基因序列ps1-dE9及App淀粉样前体蛋白基因序列APPswe融合体。设计特异PCR扩增引物,进行小鼠基因型鉴定。
ps1-dE9(NM_000021.4)的引物序列为:
上游引物如SEQ ID NO:1所述,即:5'-GAC TGA CCA CTC GAC CAG GTT CTG-3';
下游引物如SEQ ID NO:2所述,即:5'-CTT GTA AGT TGG ATT CTC ATA TCC G-3';
APPswe(NM_000484.4)的引物序列为:
上游引物如SEQ ID NO:3所述,即:5'-AAT AGA GAA CGG CAG GAG CA-3';
下游引物如SEQ ID NO:4所述,即:5'-GCC ATG AGG GCA CTA ATC AT-3'。
PCR反应条件为:95 ℃ 5 min;94 ℃ 30 s,60-65℃ 30 s,72 ℃ 40 s,72 ℃ 5min,35个循环;10℃保存。PCR产物行2%琼脂糖凝胶电泳,电泳完成后用Bio Rad凝胶电泳成像仪照相。
用常规PCR方法对7只小鼠进行鉴定,若同时扩增出分子量为350 bp和608 bp两条目的条带的为App-ps1小鼠,仅扩增出350 bp一条目的条带的为单纯App基因型的小鼠。图2是阿尔茨海默病模型小鼠的PCR基因鉴定结果图;编号1~7代表阿尔茨海默病模型小鼠个体;鉴定结果见图2,编号1、2、3、4、7仅扩增出一条350 bp目的条带,编号5、6同时扩增出350bp和608 bp两条目的条带。由此可知,编号5、6的个体为App-ps1小鼠,而编号1、2、3、4、7的个体为单纯App基因型的小鼠。
、免疫组化方法检测阿尔茨海默病模型小鼠App-ps1皮层β-淀粉样蛋白的表达
分别取2月龄和6月龄的阿尔茨海默病模型小鼠脑组织于4%多聚甲醛中固定48 h,乙醇梯度脱水,二甲苯透明,常规石蜡包埋,制成6 μm厚冠状面切片。切片常规脱蜡至水,微波炉修复抗原,冷却后PBS洗涤,然后3%过氧化氢灭活内源性过氧化物酶10 min,鼠源β-Amyloid一抗(1:800)孵育4℃冰箱过夜,PBS洗涤后,山羊抗鼠IgG二抗(1:50)孵育2 h,二氨基联苯胺(DAB)显色,苏木素复染,脱水,透明,封片;经DAB显色,应用图像分析系统测量每个视野内阳性细胞数。
图3是阿尔茨海默病模型小鼠App-ps1的β淀粉样蛋白的免疫组化结果图;图中(a)为2月龄的App-ps1小鼠,(b)为6月龄的App-ps1小鼠;由图3可见,与2月龄小鼠相比,6月龄小鼠的β-Amyloid显著增加。可见,随着月龄的增加,β-Amyloid在小鼠前额皮层表达明显增加。
、海马、皮层及纹状体FoxG1条件性过表达小鼠的鉴定
取8只实施例1中步骤(1)得到的子代海马、皮层及纹状体FoxG1条件性过表达小鼠,提取基因组DNA为模板进行双重PCR鉴定(对双重PCR鉴定的先后顺序不做限制),设计特异PCR扩增引物,测定子代小鼠的基因型。
FoxG1基因的引物序列为:
上游引物GFP如SEQ ID NO:5所述,即:5'- AAG GAC GAC GGC AAC TAC AAG -3';
下游引物GFP如SEQ ID NO:6所述,即: 5'-GGC GGT CAC GAA CTC CA -3';
Cre基因的引物序列为:
上游引物5-HT1B-tR1如SEQ ID NO:7所述:5’-AAG CTA AGT TCC TCG GGT ATGGAA G-3';
下游引物5-HT1B-tF21如SEQ ID NO:8所述:5‘-CTT CTT CAT CAT CTC CCT GGTGAT G-3';
上游引物Neo-3F1如SEQ ID NO:9所述:5'-TCT GAG GCG GAA AGA ACC AG-3';
下游引物Neo-3R如SEQ ID NO:10所述:5'-CTG GTT CTT TCC GCC TCA GA-3';
PCR反应条件为:95 ℃ 5 min;94 ℃ 30 s,65 ℃ 30 s,72 ℃ 35 s ,72 ℃ 5min;35个循环;4℃保存。PCR扩增产物进行2%琼脂糖凝胶电泳检测。电泳完成后用Bio Rad凝胶电泳成像仪照相。图4是繁育获得的海马、皮层及纹状体FoxG1条件性过表达小鼠的PCR基因鉴定结果图,M为DNA分子标记Marker;编号1~7代表海马、皮层及纹状体FoxG1条件性过表达小鼠个体;图中(a)是FoxG1基因的目的条带,(b)是Cre基因的目的条带;若PCR同时扩增出FoxG1基因的目的条带(378 bp)和Cre基因的目的条带(364 bp)的为FoxG1条件性过表达小鼠;仅扩增出一条420 bp条带为野生型小鼠;未扩增出FoxG1基因的目的条带(378 b)但扩增出Cre基因的目的条带364 bp的为Cre工具鼠;未扩增出Cre基因的目的条带364 bp,但扩增出FoxG1基因目的条带378 bp为FoxG1转基因小鼠。如图4所示,编号3、4、6、7同时扩增出378bp的 FoxG1目的条带和364bp的 Cre基因目的条带,其中,编号3、4、7还扩增有420bp的 Cre目的条带,可见,编号3、4、6、7的个体为FoxG1条件性过表达小鼠;编号1仅扩增出一条364bp 的目的条带,为Cre工具鼠,编号2仅扩增出一条420 bp 的目的条带,为野生型小鼠,编号5扩增出FoxG1基因的目的条带378 b和420 bp目的条带,为FoxG1转基因小鼠。
、阿尔茨海默病病变脑区FoxG1过表达小鼠的鉴定
取15只实施例1中制备的阿尔茨海默病病变脑区FoxG1过表达小鼠,提取基因组DNA为模板进行三重PCR鉴定(对三重PCR鉴定的先后顺序不做限制),测定子代小鼠的基因型。App-ps1基因的引物序列为SEQ ID NO:1~4,FoxG1基因的引物序列为SEQ ID NO:5~6,Cre基因的引物序列为SEQ ID NO:7~8;图5是繁育获得的阿尔茨海默病病变脑区FoxG1过表达小鼠的三重PCR基因鉴定结果图,M为DNA分子标记Marker;编号1~15代表阿尔茨海默病病变脑区FoxG1条件性过表达小鼠个体;图中,(a)是Ps1和App的目的条带,(b)是FoxG1的目的条带,(c)是Cre的目的条带;由图5所示,图(a)第一轮PCR结果表明,编号为1、2、4、5、6、8、9、10、11、12、15的个体同时扩增出608 bp和350 bp的目的条带,为含有App-ps1基因的阿尔茨海默病模型小鼠,对该模型小鼠进行第二轮PCR,图(b)表明编号5、9、10、11、12、15的个体扩增出378 bp的目的条带,含有FoxG1基因,对其进行第三轮PCR后图(c)表明,编号5、9、10、11、12、15的个体均扩增出364bp的Cre基因目的条带,其中,编号5、9、10、12、15的个体还同时扩增出420 bp的Cre条带,可见,编号5、9、10、11、12、15对应的个体为阿尔茨海默病病变脑区FoxG1过表达小鼠。
实施例三:他莫昔芬诱导阿尔茨海默病病变脑区FoxG1过表达小鼠皮层FoxG1过表达
本实施例中取6月龄的阿尔茨海默病病变脑区FoxG1过表达小鼠皮下注射他莫昔芬诱导Cre重组酶的表达,野生型小鼠作为对照,检验小鼠皮层FoxG1过表达状况。
将阿尔茨海默病病变脑区FoxG1过表达小鼠通过扩增繁殖,获得一定数量的子代至月龄6个月,小鼠腹腔注射浓度为20 mg / mL的他莫昔芬/玉米油溶液(75 mg/kg),隔天注射一次,连续两周。取小鼠脑组织于4%多聚甲醛中固定48 h后转移入30%的蔗糖溶液中,直至脑组织沉底,进行OCT包埋,-80 ℃冰箱速冻,冰冻切片机冠状面切片20 um厚。PBS冲洗30 min后,微波炉抗原修复,降至室温后PBS洗3次,10%山羊血清封闭90 min,GFP一抗(1:50)过夜。山羊抗兔IgG二抗(1:500 )避光孵育2h后,Dapi染核,甘油封片,镜检。图6是野生型小鼠与阿尔茨海默病病变脑区FoxG1过表达小鼠前额皮层FoxG1表达GFP荧光鉴定结果对比图;图中,A是野生型小鼠,B是阿尔茨海默病病变脑区FoxG1过表达小鼠;如图6可见,免疫荧光后阿尔茨海默病病变脑区FoxG1过表达小鼠与野生型小鼠相比,阿尔茨海默病FoxG1条件性过表达小鼠的前额皮层FoxG1表达具有显著增加。
以上显示和描述了本发明的基本原理、主要特征以及本发明的优点。本行业的技术人员应该了解,本发明不受上述实施例的限制,上述实施例和说明书中描述的只是说明本发明的原理,在不脱离本发明精神和范围的前提下,本发明还会有各种变化和改进,这些变化和改进都落入要求保护的本发明范围内。
序列表
<110> 镇江杰胜瑞科技有限公司
<120> 阿尔茨海默病病变区FoxG1过表达小鼠模型的构建方法及应用
<160> 10
<170> SIPOSequenceListing 1.0
<210> 1
<211> 24
<212> DNA
<213> 人工序列(Artificial Sequence)
<400> 1
gactgaccac tcgaccaggt tctg 24
<210> 2
<211> 25
<212> DNA
<213> 人工序列(Artificial Sequence)
<400> 2
cttgtaagtt ggattctcat atccg 25
<210> 3
<211> 20
<212> DNA
<213> 人工序列(Artificial Sequence)
<400> 3
aatagagaac ggcaggagca 20
<210> 4
<211> 20
<212> DNA
<213> 人工序列(Artificial Sequence)
<400> 4
gccatgaggg cactaatcat 20
<210> 5
<211> 21
<212> DNA
<213> 人工序列(Artificial Sequence)
<400> 5
aaggacgacg gcaactacaa g 21
<210> 6
<211> 17
<212> DNA
<213> 人工序列(Artificial Sequence)
<400> 6
ggcggtcacg aactcca 17
<210> 7
<211> 25
<212> DNA
<213> 人工序列(Artificial Sequence)
<400> 7
aagctaagtt cctcgggtat ggaag 25
<210> 8
<211> 25
<212> DNA
<213> 人工序列(Artificial Sequence)
<400> 8
cttcttcatc atctccctgg tgatg 25
<210> 9
<211> 20
<212> DNA
<213> 人工序列(Artificial Sequence)
<400> 9
tctgaggcgg aaagaaccag 20
<210> 10
<211> 20
<212> DNA
<213> 人工序列(Artificial Sequence)
<400> 10
ctggttcttt ccgcctcaga 20
序列表
<110> 镇江杰胜瑞科技有限公司
<120> 阿尔茨海默病病变区FoxG1过表达小鼠模型的构建方法及应用
<160> 10
<170> SIPOSequenceListing 1.0
<210> 1
<211> 24
<212> DNA
<213> 人工序列(Artificial Sequence)
<400> 1
gactgaccac tcgaccaggt tctg 24
<210> 2
<211> 25
<212> DNA
<213> 人工序列(Artificial Sequence)
<400> 2
cttgtaagtt ggattctcat atccg 25
<210> 3
<211> 20
<212> DNA
<213> 人工序列(Artificial Sequence)
<400> 3
aatagagaac ggcaggagca 20
<210> 4
<211> 20
<212> DNA
<213> 人工序列(Artificial Sequence)
<400> 4
gccatgaggg cactaatcat 20
<210> 5
<211> 21
<212> DNA
<213> 人工序列(Artificial Sequence)
<400> 5
aaggacgacg gcaactacaa g 21
<210> 6
<211> 17
<212> DNA
<213> 人工序列(Artificial Sequence)
<400> 6
ggcggtcacg aactcca 17
<210> 7
<211> 25
<212> DNA
<213> 人工序列(Artificial Sequence)
<400> 7
aagctaagtt cctcgggtat ggaag 25
<210> 8
<211> 25
<212> DNA
<213> 人工序列(Artificial Sequence)
<400> 8
cttcttcatc atctccctgg tgatg 25
<210> 9
<211> 20
<212> DNA
<213> 人工序列(Artificial Sequence)
<400> 9
tctgaggcgg aaagaaccag 20
<210> 10
<211> 20
<212> DNA
<213> 人工序列(Artificial Sequence)
<400> 10
ctggttcttt ccgcctcaga 20
Claims (8)
1.一种阿尔茨海默病病变脑区FoxG1过表达小鼠模型,其特征在于,通过5-HT1B启动子驱动Cre重组酶在海马、皮层及纹状体特定脑区特异性表达,选择性敲除FoxG1转基因小鼠CAG-loxp-stop-loxp-FoxG1-IRES-EGFP中两个loxp基因中间的stop位点,诱导App-ps1小鼠海马、皮层及纹状体FoxG1条件性过表达,构建阿尔茨海默病病变脑区FoxG1过表达小鼠。
2.一种阿尔茨海默病病变脑区FoxG1过表达小鼠模型的构建方法,其特征在于,包括如下步骤:
(1)FoxG1转基因小鼠与Cre工具小鼠交配繁育后获得海马、皮层及纹状体FoxG1条件性过表达小鼠;
(2)将繁育获得的海马、皮层及纹状体FoxG1条件性过表达小鼠与阿尔茨海默病模型小鼠交配,迅速扩大种群,得到阿尔茨海默病病变脑区FoxG1过表达小鼠;(3)他莫昔芬/玉米油溶液诱导阿尔茨海默病病变脑区FoxG1过表达小鼠FoxG1蛋白过表达;根据如SEQ ID NO:5~10所示引物序列对海马、皮层及纹状体FoxG1条件性过表达小鼠进行多重PCR鉴定;根据如SEQ ID NO:1~10所示引物序列对阿尔茨海默病病变脑区FoxG1过表达小鼠进行多重PCR鉴定。
3.根据权利要求2所述的FoxG1过表达小鼠模型的构建方法,其特征在于,采用免疫荧光技术对阿尔茨海默病病变脑区FoxG1过表达小鼠进行皮层FoxG1蛋白表达含量鉴定。
4.根据权利要求2所述的FoxG1过表达小鼠模型的构建方法,其特征在于,步骤(3)中所述他莫昔芬/玉米油溶液的浓度为20 mg / mL,所述诱导方法按每kg小鼠体重75 mg的剂量隔天注射莫昔芬/玉米油溶液一次,连续两周。
5.根据权利要求2所述的FoxG1过表达小鼠模型的构建方法,其特征在于,步骤(3)中所述阿尔茨海默病病变脑区FoxG1过表达小鼠为生长至6月龄的小鼠。
6.根据权利要求3所述的FoxG1过表达小鼠模型的构建方法,其特征在于,所述皮层FoxG1蛋白表达含量鉴定中一抗为兔源GFP,二抗为山羊抗兔IgG。
7.根据权利要求2-6任意一项所述的构建方法构建的阿尔茨海默病病变脑区FoxG1过表达小鼠模型在制备或筛选阿尔茨海默病治疗药物中的应用。
8.一种筛选阿尔茨海默病治疗药物的方法,其特征在于,将阿尔茨海默病治疗备选药物施用于如权利要求2-6任意一项所述的构建方法获得的阿尔茨海默病病变脑区FoxG1过表达小鼠模型的病状部位,通过比较用药前后小鼠病变脑区FoxG1过表达量的变化,筛选出使病变脑区FoxG1过表达量减少的药物作为阿尔茨海默病治疗药物。
Priority Applications (1)
Application Number | Priority Date | Filing Date | Title |
---|---|---|---|
CN201910933563.9A CN110622921B (zh) | 2019-09-29 | 2019-09-29 | 阿尔茨海默病病变区FoxG1过表达小鼠模型的构建方法及应用 |
Applications Claiming Priority (1)
Application Number | Priority Date | Filing Date | Title |
---|---|---|---|
CN201910933563.9A CN110622921B (zh) | 2019-09-29 | 2019-09-29 | 阿尔茨海默病病变区FoxG1过表达小鼠模型的构建方法及应用 |
Publications (2)
Publication Number | Publication Date |
---|---|
CN110622921A CN110622921A (zh) | 2019-12-31 |
CN110622921B true CN110622921B (zh) | 2021-06-15 |
Family
ID=68973805
Family Applications (1)
Application Number | Title | Priority Date | Filing Date |
---|---|---|---|
CN201910933563.9A Active CN110622921B (zh) | 2019-09-29 | 2019-09-29 | 阿尔茨海默病病变区FoxG1过表达小鼠模型的构建方法及应用 |
Country Status (1)
Country | Link |
---|---|
CN (1) | CN110622921B (zh) |
Families Citing this family (1)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
CN116210645A (zh) * | 2022-10-17 | 2023-06-06 | 青岛大学 | 一种老年痴呆症小鼠模型的构建方法及其应用 |
Family Cites Families (7)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
IN2014DN06624A (zh) * | 2005-10-18 | 2015-07-10 | Univ Colorado | |
US20090165150A1 (en) * | 2007-12-21 | 2009-06-25 | Yinghui Zhou | Directed complementation with removable gene of interest |
CN105112448B (zh) * | 2015-08-21 | 2020-06-26 | 同济大学 | Stch基因敲除动物模型的构建方法及应用 |
CN107449831A (zh) * | 2016-05-30 | 2017-12-08 | 北京师范大学 | 阿尔兹海默症蛋白标志物的用途 |
KR20190039702A (ko) * | 2016-07-05 | 2019-04-15 | 더 존스 홉킨스 유니버시티 | H1 프로모터를 이용한 crispr 가이드 rna의 개선을 포함하는 조성물 및 방법 |
CN106047930B (zh) * | 2016-07-12 | 2020-05-19 | 北京百奥赛图基因生物技术有限公司 | 一种PS1基因条件性敲除flox大鼠的制备方法 |
CN106172212A (zh) * | 2016-07-25 | 2016-12-07 | 广州道瑞医药科技有限公司 | Ccm3基因敲除小鼠模型的建立方法及用途 |
-
2019
- 2019-09-29 CN CN201910933563.9A patent/CN110622921B/zh active Active
Also Published As
Publication number | Publication date |
---|---|
CN110622921A (zh) | 2019-12-31 |
Similar Documents
Publication | Publication Date | Title |
---|---|---|
Hatini et al. | Essential role of stromal mesenchyme in kidney morphogenesis revealed by targeted disruption of Winged Helix transcription factor BF-2. | |
Schuchardt et al. | Defects in the kidney and enteric nervous system of mice lacking the tyrosine kinase receptor Ret | |
CN108029638B (zh) | 一种视网膜色素变性疾病动物模型的构建方法及应用 | |
Inoue et al. | Mouse Zic5 deficiency results in neural tube defects and hypoplasia of cephalic neural crest derivatives | |
Dennis et al. | Mutations in Hedgehog acyltransferase (Hhat) perturb Hedgehog signaling, resulting in severe acrania-holoprosencephaly-agnathia craniofacial defects | |
Sun et al. | A human YAC transgene rescues craniofacial and neural tube development in PDGFR α knockout mice and uncovers a role for PDGFRα in prenatal lung growth | |
Hims et al. | A humanized IKBKAP transgenic mouse models a tissue-specific human splicing defect | |
CN113957074B (zh) | 一种小脑共济失调疾病模型的构建方法及应用 | |
CN110622921B (zh) | 阿尔茨海默病病变区FoxG1过表达小鼠模型的构建方法及应用 | |
Chakrabarti et al. | The zinc-binding domain of Nna1 is required to prevent retinal photoreceptor loss and cerebellar ataxia in Purkinje cell degeneration (pcd) mice | |
CN104046644A (zh) | 人源化小鼠模型的构建方法和应用 | |
CN110218743B (zh) | 一种基于CRISPR/Cas9技术的牛磺酸转运体基因敲除大鼠模型的构建方法 | |
JP5119430B2 (ja) | 多発性嚢胞腎疾患関連遺伝子及びその利用 | |
Kido et al. | Expression of the human TSPY gene in the brains of transgenic mice suggests a potential role of this Y chromosome gene in neural functions | |
JP5493118B2 (ja) | 多発性嚢胞腎疾患関連遺伝子及びその利用 | |
Délot et al. | The BMP-related protein radar: a maintenance factor for dorsal neuroectoderm cells? | |
CN113558011A (zh) | 基于γ-分泌酶激活蛋白基因的干燥综合征动物模型的建立方法 | |
US20210144976A1 (en) | Animal model for the retinal degeneration by targeted pde6b gene mutation and the preparation method thereof | |
CN115125273B (zh) | 一种乳头型颅咽管瘤动物模型的构建方法及应用 | |
CN116103292B (zh) | Zdhhc21基因突变动物模型的构建方法及其应用 | |
CN117230077B (zh) | Hakai基因在RP疾病模型构建中的应用及构建方法 | |
Schmouth et al. | Non-coding-regulatory regions of human brain genes delineated by bacterial artificial chromosome knock-in mice | |
RU2815936C1 (ru) | Способ получения мышиной модели для изучения миодистрофии Дюшенна и вариантов ее терапии | |
Jansen | Generation and functional analysis of the ALS associated HNRNPA zebrafish mutants | |
CA2529633A1 (fr) | Methodes de detection de la maladie d'alzheimer |
Legal Events
Date | Code | Title | Description |
---|---|---|---|
PB01 | Publication | ||
PB01 | Publication | ||
SE01 | Entry into force of request for substantive examination | ||
SE01 | Entry into force of request for substantive examination | ||
TA01 | Transfer of patent application right | ||
TA01 | Transfer of patent application right |
Effective date of registration: 20210425 Address after: Zhenjiang City, Jiangsu Province, 212013 Jingkou District Road No. 301 Applicant after: JIANGSU University Applicant after: Zhenjiang jieshengrui Technology Co.,Ltd. Address before: 212004 building B14, R & D zone, National University Science Park Center, Zhenjiang New District, Jiangsu Province Applicant before: Zhenjiang jieshengrui Technology Co.,Ltd. |
|
GR01 | Patent grant | ||
GR01 | Patent grant |