[go: up one dir, main page]

WO2007083367A1 - Mammifère non humain capable d'exprimer un gène rapporteur spécifique à un neurone inhibiteur - Google Patents

Mammifère non humain capable d'exprimer un gène rapporteur spécifique à un neurone inhibiteur Download PDF

Info

Publication number
WO2007083367A1
WO2007083367A1 PCT/JP2006/300620 JP2006300620W WO2007083367A1 WO 2007083367 A1 WO2007083367 A1 WO 2007083367A1 JP 2006300620 W JP2006300620 W JP 2006300620W WO 2007083367 A1 WO2007083367 A1 WO 2007083367A1
Authority
WO
WIPO (PCT)
Prior art keywords
gene
vgat
human mammal
reporter gene
euron
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.)
Ceased
Application number
PCT/JP2006/300620
Other languages
English (en)
Japanese (ja)
Inventor
Yuchio Yanagawa
Yasuo Kawaguchi
Masumi Hirabayashi
Current Assignee (The listed assignees may be inaccurate. Google has not performed a legal analysis and makes no representation or warranty as to the accuracy of the list.)
Gunma University NUC
National Institute of Natural Sciences
Original Assignee
Gunma University NUC
National Institute of Natural Sciences
Priority date (The priority date 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 date listed.)
Filing date
Publication date
Application filed by Gunma University NUC, National Institute of Natural Sciences filed Critical Gunma University NUC
Priority to PCT/JP2006/300620 priority Critical patent/WO2007083367A1/fr
Publication of WO2007083367A1 publication Critical patent/WO2007083367A1/fr
Anticipated expiration legal-status Critical
Ceased legal-status Critical Current

Links

Classifications

    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
    • G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
    • G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
    • G01N33/53—Immunoassay; Biospecific binding assay; Materials therefor
    • G01N33/566—Immunoassay; Biospecific binding assay; Materials therefor using specific carrier or receptor proteins as ligand binding reagents where possible specific carrier or receptor proteins are classified with their target compounds
    • 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
    • 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
    • C12N15/63—Introduction of foreign genetic material using vectors; Vectors; Use of hosts therefor; Regulation of expression
    • C12N15/79—Vectors or expression systems specially adapted for eukaryotic hosts
    • C12N15/85—Vectors or expression systems specially adapted for eukaryotic hosts for animal cells
    • C12N15/8509—Vectors or expression systems specially adapted for eukaryotic hosts for animal cells for producing genetically modified animals, e.g. transgenic
    • 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
    • A01K2217/00—Genetically modified animals
    • A01K2217/05—Animals comprising random inserted nucleic acids (transgenic)
    • 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
    • A01K2267/0393—Animal model comprising a reporter system for screening tests
    • 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
    • C12N2800/00—Nucleic acids vectors
    • C12N2800/20—Pseudochromosomes, minichrosomosomes
    • C12N2800/204—Pseudochromosomes, minichrosomosomes of bacterial origin, e.g. BAC
    • 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
    • C12N2830/00—Vector systems having a special element relevant for transcription
    • C12N2830/008—Vector systems having a special element relevant for transcription cell type or tissue specific enhancer/promoter combination
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N2800/00—Detection or diagnosis of diseases
    • G01N2800/30—Psychoses; Psychiatry

Definitions

  • Non-human mammals that express reporter genes specifically for inhibitory neurons
  • the present invention relates to a non-human mammal that expresses a reporter gene such as a fluorescent protein gene in a suppressive-euron-specific manner.
  • the brain is made up of a collection of neural networks composed of excitability-euron and inhibitory-euron! Inhibitory-Euron's main neurotransmitters (inhibitory neurotransmitters) are ⁇ -aminobutyric acid (GABA) and glycine. Release GABA as a neurotransmitter Nerve cells are GABA neurons, which are widely distributed in the brain and spinal cord. On the other hand, neurons that release dalysin are glycine-euron and are distributed mainly in the brain stem and spinal cord. Inhibitor transmitters are known to control neuronal potential activity and play a central role in building brain functions such as arousal, sleep, circadian rhythm and learning, movement, and sensory information processing. ! / GABA has been reported to be associated with neuropsychiatric disorders such as epilepsy and alcohol psychosis, and mental symptoms such as anxiety and depression. On the other hand, glycine is known to be associated with startle disease.
  • GABA ⁇ -aminobutyric acid
  • GABA and glycine are accumulated in synaptic vesicles by the vesicular GABA transporter (VGAT) and then released into the synaptic cleft. Since VGAT is specifically expressed in inhibitory-euron (GABA and europium and glycine-euron), VGAT is also used as one of the most effective inhibitory-euron. On the other hand, inhibitory-eurone is generally scattered in the central nervous system and is relatively small in number, so it was difficult to accurately identify it in the living body and to analyze its functions such as electrophysiological characteristics.
  • Inhibitory-Green fluorescent protein gene is a glutamic acid that visualizes euron.
  • a transgenic animal having been inserted into the same reading frame as the exon of the minate decarboxylase gene has been reported (Patent Document 1).
  • Patent Document 2 An animal having a gene encoding a fluorescent protein linked so as to be under the control of the expression control region of the VGAT gene has also been reported (Patent Document 2).
  • Patent Document 2 a region of about 8 kbp of the 5 ′ expression regulatory region was used as the expression control region of the VGAT gene.
  • Patent Document 1 Japanese Patent Laid-Open No. 2003-088272
  • Patent Document 2 Japanese Patent Laid-Open No. 2003-204796
  • An object of the present invention is to provide an animal capable of efficiently observing and analyzing inhibitory-euron.
  • the present inventors diligently studied to solve the above problems. As a result, by using a non-human mammal in which a reporter gene operably linked to a DNA fragment containing a 5 'expression regulatory region of about 10 LOOkbp of the VGAT gene is retained on the chromosome, it is The inventors have found that Euron can be observed and analyzed efficiently, and have completed the present invention.
  • the present invention is as follows.
  • a non-human mammal that retains on a chromosome a reporter gene operably linked to a DNA fragment containing a 5 'expression regulatory region of about 10 OOkbp of the vesicular GABA transporter gene.
  • reporter gene is a gene in which the 3 ′ expression regulatory region of the vesicular GABA transporter gene is further linked to the 3 ′ side thereof.
  • FIG. 1 is an explanatory diagram of a BAC clone containing a VGAT gene (wild-type BAC), a targeting construct for homologous recombination, and a BAC clone modified by homologous recombination (modified BAC).
  • FIG. 2 is a diagram (photograph) showing the results of observation of the brain of a VGAT-venus rat with a fluorescence microscope.
  • FIG. 3 Diagram showing the results of immunostaining of VGAT-venus rat cerebral cortex specimens with anti-GFP antibody
  • Upper part is brain membrane and layer I
  • middle part is II ⁇ V layer
  • lower part is VI layer and white matter.
  • FIG. 4 Diagram showing the results of VGAT- venUS rat cerebellar cortex specimens observed with a fluorescence microscope (photograph)
  • the upper part is a molecular layer
  • the central part is a Purkinje cell layer
  • the lower part is a granular layer.
  • the non-human mammal of the present invention has a reporter gene chromosomally linked to a DNA fragment containing a 5 'expression regulatory region of about 1 OOkbp of the vesicular GABA transporter (VGAT) gene. A non-human mammal that is retained.
  • VGAT vesicular GABA transporter
  • the VGAT gene is not particularly limited as long as it encodes VGAT, but a mouse-derived gene that prefers a mouse or rat-derived gene is more preferable.
  • Examples of the 5 ′ expression regulatory region of about lOOkbp of the VGAT gene include a region of about lOOkbp 5 ′ to the start codon ATG contained in exon 1 of the VGAT gene.
  • the above region may be one in which one or a plurality of bases are substituted, deleted, inserted, etc., as long as the region induces reporter gene repression—specific expression in euron.
  • the term “multiple” refers to, for example, 2 to 100, preferably 2 to 50, and more preferably 2 to 20.
  • about lOOkbp is 90-: L 10 kbp, preferably 95-105 kbp, particularly preferably 99-: LOlkbp.
  • a region on the 5 ′ side may be further included.
  • the non-human mammal of the present invention retains a gene construct that is operably linked to a DNA fragment containing a 5 'expression regulatory region of about lOOkbp of the reporter gene strength VGAT gene.
  • the gene construct may be directly linked to the 5 ′ expression regulatory region, as long as the open reading frame of a single reporter gene is linked downstream of the 5 ′ expression regulatory region of the VGAT gene. , Downstream of the expression regulatory region may be linked through any sequence without inhibiting the expression of the reporter gene.
  • a gene construct in which an open reading frame of a reporter gene, preferably an open reading frame and a polyA sequence is inserted at the position of the translation initiation codon immediately after the 5 ′ expression regulatory region of the VGAT gene is desirable.
  • the gene construct contains sequences after the translation initiation codon of the VGAT gene (exons and 3 'expression control regions)! /, Or may! / ⁇ .
  • Examples of the 3 ′ expression regulatory region include a region of about 20 kbp downstream of the stop codon of the VGAT gene.
  • DNA containing the 5 'expression regulatory region of the VGAT gene is selected from a genomic library such as BAC, YAC, etc.
  • the clone containing the DNA is selected by a method commonly used, and the 5' expression regulatory region of the VGAT gene Can be obtained by cleaving with a restriction enzyme or the like, or can be obtained by polymerase chain reaction (PCR) or the like.
  • the origin of such DNA is preferably the same type as the host into which it is introduced, but is not limited to this as long as the introduced DNA can function in the host cell! /.
  • An example of a BAC library containing a 5 ′ expression regulatory region of about 10 OOkbp of the VGAT gene is clone # 160L22 available from Genome system (St. Louis, MO, U.S.A.).
  • reporter gene a light-emitting or chromogenic protein gene used for selectively visualizing inhibitory-euron, or used for selectively destroying inhibitory-euron.
  • Toxin genes that are present.
  • GFP green fluorescent protein
  • Prasher, DC et al.
  • Gene, 111, 229-233 (1992) a force that changes from green to red when irradiated with ultraviolet rays.
  • GenBank Accession No. AB085641 a gene encoding a chameleon that emits light depending on the intracellular calcium concentration
  • a Venus gene (Nat Biotechnol.
  • Examples of the chromogenic protein gene include a gene encoding j8-galactosidase.
  • Examples of the gene encoding the toxin include a gene encoding diphtheria toxin.
  • DNA fragments containing these reporter genes can be obtained by amplification by PCR or the like, or commercially available products can be used.
  • the reporter gene can be ligated to the 3 'side of the DNA containing the 5' expression regulatory region of the VGAT gene using a restriction enzyme or the like.
  • a gene construct in which a reporter gene is inserted at the start codon position of the VGAT gene can also be prepared by homologous recombination using a gene construct in which a reporter gene is inserted between sequences before and after the start codon of the VGAT gene. . Since the gene construct has a length of lOOkbp or more, it is preferable to introduce it using BAC (E. coli artificial chromosome).
  • the non-human mammal is more preferably a rat that is preferred for rodents such as mice or rats.
  • rodents such as mice or rats.
  • Use of monoclonal antibodies is difficult in mice but possible in rats, and histological analysis is easy.
  • Many physiological experiments have been performed in rats and it is easy to collate the data.
  • Analysis of small brain nuclei in the brainstem is easier in rats than in mice. Particularly preferred.
  • the gene construct prepared as described above is introduced into germ cells of a non-human mammal to produce a genetically modified animal cell, and further this is generated to produce a non-human mammal. it can.
  • Examples of the method for introducing the gene construct into the host include a method of inserting a plurality of copies of the construct into an unspecified position of the genomic DNA of the host, a method of introducing the construct into a specific position of the host genome, and the like.
  • fertilized eggs before cleavage are preferably used. like this A fertilized egg is obtained by mating a male and female of a non-human mammal.
  • fertilized eggs can be obtained by natural mating, a method of mating with males after artificially adjusting the female sexual cycle of animals is preferred.
  • follicle stimulating hormone pregnant mare serum gonadotropin; PMSG
  • luteinizing hormone human ciliary gonadotropin; hCG
  • the dose and interval of administration of these hormones should be determined appropriately according to the type of animal.
  • fertilized eggs can be obtained by administering hormonal administration to female animals and allowing them to superovulate and extracting the oviduct force on the first day after mating.
  • the genetically modified product (transgenic animal) is injected into the resulting fertilized egg by microinjection, etc., and artificially transplanted into the female oviduct and implanted to give birth. Can be obtained.
  • LHRH luteinizing hormone-releasing hormone
  • a method in which a fertilized egg is artificially transplanted and implanted in a pseudopregnant female animal may be used.
  • the selection of pups or pups introduced with the above gene construct is performed by cutting the tip of the mouse or rat tail and extracting the polymer DNA (supervised by Tatsuji Nomura Genomic DNA is extracted by using a commercially available kit such as Motoya Katsuki, Kodansha (1987)) or DNAeasy Tissue Kit (manufactured by QIAGEN).
  • the reporter gene in the VGAT expression control region can be confirmed by confirming the presence of DNA.
  • the reporter gene is a fluorescent protein gene, it can also be measured by measuring the amount of fluorescence emitted or observing with a fluorescence microscope or the like.
  • the non-human mammal of the present invention is bred in an ordinary breeding environment after mating the individuals obtained as described above and confirming that the introduced DNA is stably maintained.
  • the progeny can be obtained by this method, or the offspring can be obtained by repeating in vitro fertilization to maintain the strain.
  • non-human mammal of the present invention retains the above gene construct on the chromosome, other It may be an animal obtained by crossing with other animals.
  • crossing the rat of the present invention with an epilepsy model animal such as an SHR rat allows observation of the inhibitory-euron dynamics in epilepsy, It is possible to obtain animals useful for analysis of the etiology of epilepsy and screening for therapeutic drugs.
  • VGAT is also known to be expressed on splenic islets of Langernons.
  • rats of this invention with diabetes model animals such as GK rats, Animals useful for observation, analysis of the etiology of diabetes, and screening of therapeutic agents can be obtained.
  • the non-human gene-modified animal of the present invention is an inhibitor of GABA neurons and the like, electrophysiological analysis and morphology of GABA neurons using the amount of protein translated by the reporter gene as an index. It can be used for scientific analysis. It can also be used for the developmental analysis of GABA-euron, molecular biological analysis specific to GABA-euron, and in vivo expression dynamics analysis of VGAT gene.
  • the expression level of the fluorescent protein can be observed and measured as the fluorescence amount.
  • the method for measuring the amount of fluorescence can be visualized by a fluorescence microscope or the like, quantified by FRET (Fluorescence Resonance Energy Transfer), the two-photon excitation method, or flow cytometry.
  • the inhibitory euron derived from the animal of the present invention is electrically or drug-stimulated, and this stimulation changes the fluorescence intensity in inhibitory neurons or synapses.
  • this stimulation changes the fluorescence intensity in inhibitory neurons or synapses.
  • functional or morphological changes of nerve cells or synapses to the stimulation can be analyzed.
  • Such an analysis elucidates the mechanism of action and interaction of physiological pharmacological effects induced by drugs or stimuli on inhibitory neurons or synapses in the central nervous system. The knowledge gained from these analyzes will lead to the elucidation of the mechanisms of neurotransmission and pharmacological action.
  • the non-human genetically modified animal of the present invention can visualize inhibitory neurons or synapses in a living body, it can be used for screening and evaluation of neurotransmitters in nerve cells or synapses. it can.
  • the non-human mammal of the present invention is used to treat the animal with a test substance, analyze changes in nerve cells or synapses in the animal, and treat this with an untreated control animal. This can be done by comparing with
  • test substance examples include peptides, proteins, non-peptidic compounds, synthetic compounds, fermentation products, cell extracts, plant extracts, animal tissue extracts, etc., and these substances are novel substances. It may be a known substance. Examples of a method for treating the animal with a test substance include known methods commonly used such as oral administration and intravenous injection, and can be appropriately selected according to the symptom of the test animal, the nature of the test substance, and the like. Good.
  • screening method of the present invention can also be performed using nerve tissue obtained from the genetically modified animal of the present invention.
  • Examples of changes that appear in nerve cells or synapses include structural or morphological effects that accompany changes in the expression level of fluorescent proteins and the number of synapses.
  • BAC E. coli artificial chromosome
  • the # 160L22 clone has about 100 kb on the 5 ′ side of the VGAT gene, about 25 kb on the 3 ′ side, and has an insert of about 130 kb in total, including the VGAT gene (about 5 kb). found.
  • the s gene (Nat Biotechnol. 2002 Jan; 20 (l): 87-90.) was inserted. In other words, it contains the Venus gene followed by the SV40poly (A) sequence, its 5-end is the VGAT gene 5-homology region (0.94 kb 5 '-expression control region), and the 3'-end is VGAT gene A DNA containing a 3'-homologous region (a region containing 1.39 kb of Ettason 1 and its downstream region) was prepared and used as a homologous recombination shuttle vector (pKOV—Kan; Lalioti and Heath, Nucleic Acids Res.
  • pKOV homologous recombination shuttle vector
  • VGAT-venus / KOV a targeting construct, VGAT-venus / KOV (Fig. 1) was prepared.
  • VGAT-venus / KOV was introduced into # 160L2 2 clones, and by homologous recombination, VGAT-venus BAC clone with venus gene and poly A sequence inserted into ATG position of VGAT gene on # 160L22 clone was Lalioti and It was prepared according to the method of Heath (2001).
  • BAC DNA was purified from the VGAT-venus BAC clone prepared as described above, and the DNA was injected into a rat fertilized egg and then transplanted into a pseudopregnant mouse to obtain 21 offspring ( founder). Whether or not the VGAT-venus gene was introduced was determined by PCR. The prime r used was designed in the venus gene, and the sequence is as follows.
  • venus- R 5,-TTACTTGTACAGCTCGTCCATGCCGA-3 '(SEQ ID NO: 2).
  • FIG. 3 shows the results using cerebral cortex specimens. Anti-GFP antibody-positive cells are small to medium in size, have various morphologies, and have scattered GABA-euron characteristics. In addition, using GFP autofluorescence and anti-GABA antibody In addition, double staining revealed that GFP autofluorescent cells were anti-GABA antibody positive cells.
  • Figure 4 shows the results using cerebellar cortex specimens.
  • the non-human mammal of the present invention is an inhibitory-euron that efficiently expresses a reporter gene. Therefore, by using the non-human mammal of the present invention, inhibitory-euron can be clearly identified and visualized under living conditions. In addition, electrophysiological experiments for analyzing the molecular mechanism of information transmission and pharmacological experiments for investigating the activity of various drugs acting on inhibitory neurons and their mechanism of action are much easier than before. Can be done. By using the nerve cells or synapses of the non-human mammal of the present invention, screening for therapeutic agents for cranial nerve diseases such as anxiety, depression, epilepsy, and nerve cell death can be performed efficiently.
  • cranial nerve diseases such as anxiety, depression, epilepsy, and nerve cell death

Landscapes

  • Life Sciences & Earth Sciences (AREA)
  • Health & Medical Sciences (AREA)
  • Engineering & Computer Science (AREA)
  • Chemical & Material Sciences (AREA)
  • Biomedical Technology (AREA)
  • Biotechnology (AREA)
  • Genetics & Genomics (AREA)
  • Immunology (AREA)
  • Zoology (AREA)
  • General Health & Medical Sciences (AREA)
  • Molecular Biology (AREA)
  • Biochemistry (AREA)
  • Hematology (AREA)
  • Organic Chemistry (AREA)
  • Physics & Mathematics (AREA)
  • Wood Science & Technology (AREA)
  • Urology & Nephrology (AREA)
  • Microbiology (AREA)
  • Bioinformatics & Cheminformatics (AREA)
  • General Engineering & Computer Science (AREA)
  • Veterinary Medicine (AREA)
  • Environmental Sciences (AREA)
  • Pathology (AREA)
  • Animal Husbandry (AREA)
  • Biodiversity & Conservation Biology (AREA)
  • Animal Behavior & Ethology (AREA)
  • Cell Biology (AREA)
  • Biophysics (AREA)
  • Food Science & Technology (AREA)
  • Plant Pathology (AREA)
  • General Physics & Mathematics (AREA)
  • Analytical Chemistry (AREA)
  • Medicinal Chemistry (AREA)
  • Micro-Organisms Or Cultivation Processes Thereof (AREA)
  • Measuring Or Testing Involving Enzymes Or Micro-Organisms (AREA)

Abstract

L'invention concerne un animal permettant une observation et une analyse efficaces d'un neurone inhibiteur grâce à la rétention, sur un chromosome d'un mammifère non humain, d'un gène rapporteur lié de manière exprimable à un fragment d'ADN qui contient une région régulant l'expression en 5’ d'environ 100 kbp du gène codant pour les transporteurs vésiculaires du GABA.
PCT/JP2006/300620 2006-01-18 2006-01-18 Mammifère non humain capable d'exprimer un gène rapporteur spécifique à un neurone inhibiteur Ceased WO2007083367A1 (fr)

Priority Applications (1)

Application Number Priority Date Filing Date Title
PCT/JP2006/300620 WO2007083367A1 (fr) 2006-01-18 2006-01-18 Mammifère non humain capable d'exprimer un gène rapporteur spécifique à un neurone inhibiteur

Applications Claiming Priority (1)

Application Number Priority Date Filing Date Title
PCT/JP2006/300620 WO2007083367A1 (fr) 2006-01-18 2006-01-18 Mammifère non humain capable d'exprimer un gène rapporteur spécifique à un neurone inhibiteur

Publications (1)

Publication Number Publication Date
WO2007083367A1 true WO2007083367A1 (fr) 2007-07-26

Family

ID=38287329

Family Applications (1)

Application Number Title Priority Date Filing Date
PCT/JP2006/300620 Ceased WO2007083367A1 (fr) 2006-01-18 2006-01-18 Mammifère non humain capable d'exprimer un gène rapporteur spécifique à un neurone inhibiteur

Country Status (1)

Country Link
WO (1) WO2007083367A1 (fr)

Citations (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
JP2003204796A (ja) * 2001-11-08 2003-07-22 Japan Science & Technology Corp 非ヒト遺伝子改変動物およびその利用

Patent Citations (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
JP2003204796A (ja) * 2001-11-08 2003-07-22 Japan Science & Technology Corp 非ヒト遺伝子改変動物およびその利用

Non-Patent Citations (4)

* Cited by examiner, † Cited by third party
Title
CHIU C.-S. ET AL.: "Number, density, and surface/cytoplasmic distribution of GABA transporters at presynaptic structures of knock-in mice carrying GABA transporter subtype 1-green fluorescent protein fusions", J. NEUROSCI., vol. 22, no. 23, 1 December 2002 (2002-12-01), pages 10251 - 10266, XP002996448 *
EBIHARA S. ET AL.: "Mouse vesicular GABA transporter gene: geomic organization, transcriptional regulation and chromosomal localization", MOL. BRAIN RES., vol. 110, 2003, pages 126 - 139, XP002997402 *
OH W-J. ET AL.: "The mouse vesicular inhibitory amino acid gene: Expression during embryogenesis, analysis of its core promoter in neural stem cells and a reconsideration of its altenate splicing", GENE, vol. 351, 23 May 2005 (2005-05-23), pages 39 - 49, XP049388396 *
SAGNE C. ET AL.: "Cloning of a functional vesicular GABA and glycine transporter by screening of genome databases", FEBS LETT., vol. 417, no. 2, 10 November 1997 (1997-11-10), pages 177 - 183, XP002916396 *

Similar Documents

Publication Publication Date Title
Lin et al. Conditional deletion of hippocampal CA2/CA3a oxytocin receptors impairs the persistence of long-term social recognition memory in mice
Qian et al. Sleep homeostasis regulated by 5HT2b receptor in a small subset of neurons in the dorsal fan-shaped body of drosophila
Lee et al. Genetic and neuronal regulation of sleep by neuropeptide VF
Zhang et al. Rapid in vivo functional analysis of transgenes in mice using whole body imaging of luciferase expression
Hirrlinger et al. Expression of reef coral fluorescent proteins in the central nervous system of transgenic mice
Horie et al. Investigation of Oxtr-expressing neurons projecting to nucleus accumbens using Oxtr-ires-Cre knock-in prairie voles (Microtus ochrogaster)
JPWO2003041496A1 (ja) トランスジェニック動物
Homma et al. KIF2A regulates the development of dentate granule cells and postnatal hippocampal wiring
Mitsui et al. Genetic visualization of the secondary olfactory pathway in Tbx21 transgenic mice
Nelson et al. Molecular and circuit architecture of social hierarchy
TW200529742A (en) Method for making a non-human mammal
CN104975042A (zh) 在髓鞘碱性蛋白启动子的控制下表达荧光素酶的动物模型及其用途
JP4872072B2 (ja) 抑制性ニューロン特異的にレポーター遺伝子を発現する非ヒト哺乳動物
US8952213B2 (en) Neuronal activation in a transgenic model
JP2003204796A (ja) 非ヒト遺伝子改変動物およびその利用
WO2007083367A1 (fr) Mammifère non humain capable d'exprimer un gène rapporteur spécifique à un neurone inhibiteur
AU2003285573A1 (en) Fish disease models and uses thereof
Yu et al. Advantages and differences among various animal models of Huntington’s disease
Perry et al. Fish physiology: zebrafish
US20120036588A1 (en) Method for observing gad67-positive cell in transgenic animal
JPWO2001033957A1 (ja) 細胞内のカルシウムイオンの濃度変化をモニターするトランスジェニック非ヒト哺乳動物
Balu et al. Behavioral and physiological characterization of PKC-dependent phosphorylation in the Grin2a∆ PKC mouse
KR100517831B1 (ko) ＰＳ２ 돌연변이 유전자 및 ＡＰＰｓｗ 유전자를 발현하는 이중 형질전환 치매 마우스
Ikpatt An analysis of gene expression in the socially monogamous brain
JP3949520B2 (ja) レギュカルチン過剰発現モデル動物

Legal Events

Date Code Title Description
121 Ep: the epo has been informed by wipo that ep was designated in this application
DPE1 Request for preliminary examination filed after expiration of 19th month from priority date (pct application filed from 20040101)
NENP Non-entry into the national phase

Ref country code: DE

122 Ep: pct application non-entry in european phase

Ref document number: 06711889

Country of ref document: EP

Kind code of ref document: A1

NENP Non-entry into the national phase

Ref country code: JP