[go: up one dir, main page]

WO2006085590A1 - ANIMAL TRANSGÉNIQUE NON HUMAIN EXPRIMANT LE TetR ET PROCÉDÉ SERVANT À PRODUIRE CELUI-CI - Google Patents

ANIMAL TRANSGÉNIQUE NON HUMAIN EXPRIMANT LE TetR ET PROCÉDÉ SERVANT À PRODUIRE CELUI-CI Download PDF

Info

Publication number
WO2006085590A1
WO2006085590A1 PCT/JP2006/302265 JP2006302265W WO2006085590A1 WO 2006085590 A1 WO2006085590 A1 WO 2006085590A1 JP 2006302265 W JP2006302265 W JP 2006302265W WO 2006085590 A1 WO2006085590 A1 WO 2006085590A1
Authority
WO
WIPO (PCT)
Prior art keywords
transgenic animal
human transgenic
tetr
expression
gene
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/JP2006/302265
Other languages
English (en)
Japanese (ja)
Inventor
Yoshitaka Tamai
Yukiko Tsukada
Naomi Murai
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.)
MSD KK
Original Assignee
Banyu Phamaceutical Co Ltd
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 Banyu Phamaceutical Co Ltd filed Critical Banyu Phamaceutical Co Ltd
Publication of WO2006085590A1 publication Critical patent/WO2006085590A1/fr
Anticipated expiration legal-status Critical
Ceased legal-status Critical Current

Links

Classifications

    • 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
    • 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
    • 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
    • A01K2267/01—Animal expressing industrially exogenous proteins
    • 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

Definitions

  • TetR-expressing non-human transgenic animal and method for producing the same
  • the present invention relates to a TetR-expressing non-human transgenic animal used for inducing expression of a desired gene in vivo.
  • Non-Patent Document 1 Science, 274, 1678-1683, 1996
  • Non-Patent Document 2 Mol. Pharmacology, 54, 495-503, 1998
  • the present invention has been made in view of the above-mentioned problems of the prior art, and is a transgene that can be used for in vivo expression induction of molecules that cannot be expressed sufficiently when induced with drugs.
  • the goal is to get a nick mouse.
  • the present inventors have used a specific promoter and introduced a sequence that contributes to high-efficiency expression.
  • the present invention was completed by successfully developing a non-human transgenic animal that can be used for inducing expression of molecules and / or proteins at a desired timing.
  • the non-human transgenic animal of the present invention is characterized by expressing a TetR protein.
  • the promoter used for expressing the TetR protein is preferably a CAG promoter.
  • the TetR gene can be expressed more strongly.
  • the TetR protein is expressed in the non-human transgenic animal of the present invention.
  • Insulator sequences are included in the expression vector. By including the insulator sequence, it becomes possible to induce the expression of the transgene with high probability.
  • an IRES sequence may be contained in an expression vector for expressing the TetR protein.
  • cells that express the TetR protein at a high level can be selected with high efficiency.
  • an expression vector in which a TetR gene is arranged downstream of a promoter is introduced into a fertilized egg or embryonic stem cell, and a TetR gene-introduced cell is obtained. And a step of introducing the TetR gene-introduced cell into a non-human mammal to obtain a TetR gene-introduced transgenic animal.
  • the method for producing a non-human transgenic animal of the present invention as a method for introducing the expression vector into a fertilized egg or embryonic stem cell, an electoral mouth location, a ribofusion, a calcium phosphate coprecipitation method is used. Any one selected from the group consisting of microinjection and viral vector methods is preferably used.
  • the promoter is a CAG promoter.
  • the TetR gene can be expressed more strongly.
  • the expression vector may further contain an insulator sequence.
  • an insulator sequence By including an insulator sequence, it is possible to induce the expression of the transgene with high probability.
  • the expression vector may contain an IRES sequence.
  • IRES sequence By including the IRES sequence, cells that express TetR protein at a high level can be selected with high efficiency.
  • test nucleic acid Tet-inducible non-human transgenic animal of the present invention is characterized by being produced by crossing the above-mentioned non-human transgenic animal and an animal expressing the test nucleic acid. .
  • Such an animal makes it possible to induce a desired test nucleic acid with tetracycline in vivo, and to analyze the function of the test nucleic acid (target gene).
  • test nucleic acid Tet-inducible non-human transgenic animal of the present invention
  • the test nucleic acid is siRNA and / or shRNA.
  • siRNA and shRNA Is a nucleic acid having a relatively small molecular weight, but by using the non-human transgenic animal of the present invention, expression can be induced in vivo by tetracycline.
  • the “promoter” in the present invention means a region on DNA that determines the transcription start site of a gene and regulates the frequency of transcription.
  • RNA interference is a phenomenon that causes double-stranded RNA force-specific gene sile ndng (gene expression suppression) that is complementary to a certain mRNA and specifically suppresses the expression of the target gene. It is used as a technology.
  • CAG promoter is a promoter which has been reported to work strongly in many cells and animals as a tissue non-specific expression promoter.
  • non-human transgenic animal that can be used when inducing expression of a molecule in vivo when sufficient expression cannot be obtained when induced with a drug.
  • it can be used to induce the expression of low molecular weight nucleic acid molecules that cannot be expressed sufficiently when induced with drugs in vivo, or to induce the expression of low expression proteins with high efficiency.
  • Non-human transgenic animals can be obtained.
  • FIG. 1 is a view showing the expression of TetR protein in each tissue confirmed by Western analysis. BEST MODE FOR CARRYING OUT THE INVENTION
  • the non-human transgenic animal of the present invention is characterized by expressing TetR protein.
  • the expression induction system by the tetracycline gene is a system that utilizes the characteristics of the Tet repressor (TetR protein) and Tet operator (tetO) sequences that act on the transcriptional regulation of the tetracycline-resistant operon of Escherichia coli.
  • TetR protein Tet repressor
  • tetO Tet operator
  • Tetracycline Nyxixa It is possible to start / stop the expression of the desired gene by induction of iclin and the like (including Tet).
  • TetTA Tet transactivator
  • rtTA reverse Tet transactivator
  • a system using tTA or rtTA often uses a pol II promoter transcribed by RNA polymerase II.
  • the promoters used in the system have insufficient expression ability to express a relatively small molecular weight nucleic acid (eg, siRNA or shRNA), and sufficient expression cannot be expected. Therefore, in producing the non-human transgenic animal of the present invention, a tetracycline-induced expression system using tTA or rtTA cannot be used.
  • the present inventors constructed a TetR expression system using a promoter that can be expected to have higher expression or a promoter of ⁇ ⁇ ⁇ ⁇ ⁇ ⁇ ⁇ system, and desired low molecular nucleic acid and Z or protein. We succeeded in producing transgenic mice that can be used for the expression of.
  • the TetR gene according to the present invention may be a mutant having a substitution, deletion, addition or insertion of one or more bases in the TetR gene as long as it encodes a functional protein as a Tet repressor. Yo! In addition, restriction enzyme sites and linkers necessary for DNA recombination can be added to the TetR gene as needed.
  • the type of promoter according to the present invention is not particularly limited.
  • the CA G promoter Niwa, H et al., Gene 107, 193-200 (1991)
  • tRNA promoter tRNA promoter
  • U6 promoter HI promoter tRNA promoter
  • a TetR gene is arranged downstream of the promoter. Even if the promoter and TetR gene are directly linked, The desired base sequence may be inserted, but it must be functionally linked.
  • the term “functionally linked” means that the promoter and the TetR gene are combined so that the expression of the TetR protein is induced with the activation of the transcription of the promoter.
  • the expression vector may contain other components necessary for gene expression, detection of the expressed gene, and the like.
  • examples of such components include an IRES (internal ribosome entry site) lj, a GFP (Green Fluorescent Protein) gene, and an insulator lj.
  • the IRES distribution IJ refers to the ribosome binding sequence in the mRNA, and it is incorporated into the expression vector so that one kind of mRNA force has two open reading frames. Translation is possible (J. Virology, 62, p2636_2643 (1988)).
  • the ribosome translates the target gene (TetR) from the 5 'end of the bicistronic mRNA and also translates the downstream selectable marker and reporter gene from the IRES sequence.
  • TetR target gene
  • cells that express the target protein at a high level can be selected with high efficiency by the action of a selection marker whose translation efficiency is increased by the IR ES sequence.
  • the GFP protein is a fluorescent protein derived from a luminescent jellyfish.
  • a fusion protein of a TetR gene and a GFP gene is constructed and incorporated into an expression vector to express the fusion protein. It emits green fluorescence in the body by irradiating the excitation wavelength. Using this fluorescence as an index, the expression of TetR protein can be detected visually.
  • the insulator sequence refers to a DNA segment that has a function of acting on a promoter to enhance or suppress gene expression and a function of avoiding the position effect of the transgene (Cell, 74, p505, (1993)).
  • the expression level of the transgene may differ depending on the location of the integration. In such a case, by sandwiching both sides of the transgene with an insulator sequence, a decrease in the expression level of the transgene due to the position effect is avoided. As a result, transgene expression can be guided with high probability.
  • the non-human transgenic animal of the present invention is not particularly limited as long as it is a non-human animal, and examples thereof include rodents such as mice, rats, guinea pigs, and rabbits. Can be mentioned.
  • an expression vector having the TetR gene as described above is constructed by recombinant DNA technology. Enzymatic construction can be carried out by methods well known to those skilled in the art. Enzymes such as restriction enzymes and ligases used for construction may be appropriately selected for the restriction enzyme site to be used.
  • the expression vector contains a positive selection marker such as a neomycin resistance gene, a hygromycin resistance gene or a puromycin resistance gene in order to select and concentrate the obtained homologous recombinants.
  • negative selection markers such as diphtheria toxin A gene or simple herpesvirus thymidine kinase gene may be included.
  • the constructed expression vector is introduced into an animal to produce a non-human transgenic animal.
  • a powerful non-human transgenic animal can be produced by a method well known to those skilled in the art.
  • an expression vector is introduced into the pronucleus of a fertilized egg, and the resulting cell is returned to the foster oviduct.
  • a method for introducing an expression vector into a fertilized egg or embryonic hepatocyte can also be carried out by a method well known to those skilled in the art.
  • electopore position, lipofuxion, calcium phosphate coprecipitation method, microinjection, and viral vector The method by is mentioned.
  • the method using the virus vector include methods using SV_40, adenovirus, herpesvirus, adeno-associated virus (AAV), vaccinia virus, ushipapilloma virus, and retrovirus as virus vectors. It is preferable to use retrowinoles (for example, lentivirus) as a viral vector.
  • the TetR protein is expressed in the tissue based on the tissue specificity of the expression of the promoter used. Therefore, power
  • the expression of the test nucleic acid can be induced by Tet administration by crossing the non-human transgenic animal with the non-human transgenic animal into which the test nucleic acid (transgene) has been introduced. Human transgenic animals can be generated.
  • the test nucleic acid is a nucleic acid having a relatively small molecular weight (siRNA or shRNA) or a low molecular weight nucleic acid that is difficult to express in a normal expression system, the expression can be induced.
  • siRNA or shRNA a nucleic acid having a relatively small molecular weight
  • shRNA a relatively small molecular weight nucleic acid that is difficult to express in a normal expression system
  • a non-human transgenic animal into which a test nucleic acid (transgene) has been introduced is an expression vector that contains a TetO sequence that is a binding sequence of TetR protein in addition to the test nucleic acid (transgene). It is necessary to have one. That is, it is necessary to have an expression vector in which a test nucleic acid (transgene) is arranged downstream of the TetO sequence that is a binding sequence of the TetR protein.
  • the test nucleic acid can be expressed in a Tet-inducible manner.
  • an expression vector for a test nucleic acid (transgene) transgene is further introduced into an egg (TetR + egg) or tissue (TetR + tissue) isolated from the non-human transgenic animal of the present invention.
  • an egg TetR + egg
  • tissue TetR + tissue
  • the non-human transgenic animal capable of expressing the test nucleic acid in a Tet-inducible manner can be used for functional analysis of the test nucleic acid (target gene), and the test nucleic acid (target gene) can be used. It is useful as a model animal for diseases involving offspring.
  • the primer contains a Pstl cleavage site and an Xhoi cleavage site.
  • G SEQ ID NO: 1
  • XhoI-globin-PCR CTCGAGGTGA GTTTGGGGAC CCTTGATTGT TC (SEQ ID NO: 2)
  • the amplified 1.3 kb fragment was cloned as a blunt end into the EcoRV site of pBluescript SK (-). From the cloned plasmid, fragments of the Rabitt beta globin intron II and TetR coding sequences were excised at the Pstl and Xhol cleavage sites.
  • pIRES-EGFP plasmid (Clontech) was cleaved at the Pstl cleavage site and the Xhol cleavage site. By ligating both, the pGlobin-TetR-IRES-EGFP plasmid was cloned.
  • pGlobin-TetR—IRES-EGFP plasmid strength excise the Globin-TetR-IRES-EGFP fragment with Xhol and Notl, and the EcoRI-cut pSK / CAGGS plasmid prepared in advance and ligated after blunt end treatment and pSK The / CAG-TetR-IRES-EGFP plasmid was cloned.
  • pSK / CAG-TetR-IRES-EGFP a 1.2 Kb insulator sequence at the No tl cleavage site located 5 'upstream of the CAG promoter and the Clal cleavage site located 3' downstream of the polyA signal sequence, respectively.
  • a pSKM / 1 ns-CAG-TetR-IRES-EGFP plasmid in which an CAG-TetR_IRES-EGFP expression cassette was sandwiched between insulator sequences was prepared.
  • the insulator sequence the sequence reported in Cell 74, pp505-514 was used.
  • the pCAGGS plasmid (Chemical and Serological Therapy Laboratory) was cleaved with EcoRI, and blunt-ended with a fragment of the TetR coding sequence after the blunt end treatment to clone the pCAG-TetR plasmid.
  • pCAG-TetR plasmid On the pCAG-TetR plasmid, two 1.2 Kb insulator sequences were cloned in tandem at the Notl cleavage site located 5 'upstream of the CAG promoter and the Clal cleavage site located 3' downstream of the polyA signal sequence, respectively. That is, a pSKM / Ins-CAG-TetR plasmid was prepared in which the CAG_TetR-IRES-EGFP expression cassette was sandwiched between insulator sequences. As the insulator sequence, the sequence reported in Cell 74, pp505-514 was used.
  • the insert and the vector were separated by electrophoresis on agarose gel after digestion with restriction enzymes as follows.
  • DNA fragment purification kit Qiagen
  • TE buffer a DNA fragment purification kit
  • the DNA solution thus obtained is filled into a glass capillary and microinjected into the female pronucleus of a fertilized egg of a mouse. Injected. Furthermore, after the fertilized egg was transplanted into the oviduct of a foster mother of pseudopregnancy, a transgenic mouse was obtained by spontaneous delivery.
  • the present invention it is possible to obtain a transgenic animal that can be used for inducing expression of a nucleic acid molecule and / or protein in vivo when sufficient expression is not obtained when induced with a drug. It becomes possible. Therefore, control the expression of the desired gene by expressing siRNA and / or shRNA against the gene of interest using a powerful non-human transgenic animal or by expressing a protein that is the translation product of the gene. And the in vivo function of the gene can be analyzed.

Landscapes

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

Abstract

L'invention a pour objet d'obtenir une souris transgénique non humaine qu'on peut utiliser pour induire l'expression in vivo d'une molécule d'acide nucléique et/ou d'une protéine qui ne sont pas exprimées suffisamment lorsqu'elles sont induites par une substance médicamenteuse ; et un procédé servant à produire celui-ci. Avec le procédé servant à produire un animal transgénique non humain caractérisé en ce qu'il comprend les étapes consistant à produire une cellule dans laquelle le gène TetR est introduit en introduisant dans un oeuf fécondé ou une cellule souche d'embryon un vecteur d'expression dans lequel le gène TetR est situé en aval d'un promoteur ; et obtenir un animal transgénique non humain dans lequel le gène TetR est introduit en introduisant chez un mammifère non humain la cellule dans laquelle le gène TetR est introduit ; et avec l'animal transgénique non humain produit par le procédé de production, il devient possible d'obtenir un animal transgénique non humain qu'on peut utiliser lorsqu'on induit in vivo l'expression d'une molécule d'acide nucléique et/ou d'une protéine qui ne sont pas exprimées suffisamment lorsqu'elles sont induites par une substance médicamenteuse.
PCT/JP2006/302265 2005-02-09 2006-02-09 ANIMAL TRANSGÉNIQUE NON HUMAIN EXPRIMANT LE TetR ET PROCÉDÉ SERVANT À PRODUIRE CELUI-CI Ceased WO2006085590A1 (fr)

Applications Claiming Priority (2)

Application Number Priority Date Filing Date Title
JP2005-032664 2005-02-09
JP2005032664A JP2008104357A (ja) 2005-02-09 2005-02-09 ＴｅｔＲ発現非ヒトトランスジェニック動物及びその製造方法

Publications (1)

Publication Number Publication Date
WO2006085590A1 true WO2006085590A1 (fr) 2006-08-17

Family

ID=36793161

Family Applications (1)

Application Number Title Priority Date Filing Date
PCT/JP2006/302265 Ceased WO2006085590A1 (fr) 2005-02-09 2006-02-09 ANIMAL TRANSGÉNIQUE NON HUMAIN EXPRIMANT LE TetR ET PROCÉDÉ SERVANT À PRODUIRE CELUI-CI

Country Status (2)

Country Link
JP (1) JP2008104357A (fr)
WO (1) WO2006085590A1 (fr)

Cited By (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US11643666B2 (en) 2017-10-25 2023-05-09 Nouscom Ag Eukaryotic cell line

Non-Patent Citations (6)

* Cited by examiner, † Cited by third party
Title
KUNINGER D. ET AL.: "Gene disruption by regulated short interfering RNA expression, using a two-adenovirus system", HUM. GENE THER., vol. 15, no. 12, 2004, pages 1287 - 1292, XP002350362 *
LU L. AND TOWER J.: "A transcriptional insulator element, the su(Hw) binding site, protects a chromosomal DNA replication origin from position effects", MOL. CELL., vol. 17, no. 4, 1997, pages 2202 - 2206, XP002999184 *
LYCETT G.J. ET AL.: "Conditional expression in the malaria mosquito Anopheles stephensi with Tet-On and Tet-Off systems", GENETICS, vol. 167, no. 4, 2004, pages 1781 - 1790, XP002999183 *
NIWA H. ET AL.: "Efficient selection for high-expression transfectants with a novel eukaryotic vector", GENE, vol. 108, no. 2, 1991, pages 193 - 199, XP000569330 *
REES S. ET AL.: "Bicistronic vector for the creation of stable mammalian cell lines that predisposes all antibiotic-resistant cells to express recombinant protein", BIOTECHNIQUES, vol. 20, no. 1, 1996, pages 102 - 104, AND 106, AND 108 - 110, XP001093737 *
SHIGEHARA T. ET AL.: "Inducible podocyte-specific gene expression in transgenic mice", J. AM. SOC. NEPHROL., vol. 14, 2003, pages 1998 - 2003, XP002999182 *

Cited By (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US11643666B2 (en) 2017-10-25 2023-05-09 Nouscom Ag Eukaryotic cell line

Also Published As

Publication number Publication date
JP2008104357A (ja) 2008-05-08

Similar Documents

Publication Publication Date Title
US10988776B2 (en) Methods of modifying genes in eukaryotic cells
JP4375759B2 (ja) 標的とされた発現特性に従う異種遺伝子の発現
EP2262898B1 (fr) Souris issues de cellules souches embryonnaires par injection d'embryons hôtes diploïdes
JP2008505625A (ja) リコンビナーゼを介したカセット交換を用いる、短いヘアピン型ｒｎａ発現カセットの標的遺伝子組換え
AU2021252114A1 (en) Methods for targeted integration
CN102669057B (zh) 一种调控动物内源基因表达的遗传修饰方法
JP2004530429A (ja) 転写活性な座を標的化する方法
US7625755B2 (en) Conditional knockout method for gene trapping and gene targeting using an inducible gene silencer
US20120107938A1 (en) Methods and kits for high efficiency engineering of conditional mouse alleles
WO2025002401A1 (fr) Animal non humain à gène cfb modifié
Kleinhammer et al. Conditional RNAi in mice
WO2006085590A1 (fr) ANIMAL TRANSGÉNIQUE NON HUMAIN EXPRIMANT LE TetR ET PROCÉDÉ SERVANT À PRODUIRE CELUI-CI
JP5481661B2 (ja) 変異導入遺伝子作製方法
JP2018526005A (ja) 前立腺がんのげっ歯類モデル
KR102131883B1 (ko) Cre 재조합효소 의존적으로 TREK-1 발현이 억제되는 항우울 동물모델 및 이의 제조방법
US20010049828A1 (en) Genetic system for controlling background expression of transgene products
CN103849634B (zh) 转基因构建物及其在制备附睾头部基因条件性敲除小鼠模型中的应用
US20080153764A1 (en) System and Methods For Short Rna Expression
JP2014103906A (ja) 部位特異的組換え系を利用した遺伝子導入方法およびそのためのキット
WO2000062604A1 (fr) Systeme genetique destine a reguler l'expression en arriere-plan de produits transgeniques
WO2006055416A2 (fr) Souris transgenique pouvant surexprimer le ve-ptp

Legal Events

Date Code Title Description
121 Ep: the epo has been informed by wipo that ep was designated in this application
NENP Non-entry into the national phase

Ref country code: DE

122 Ep: pct application non-entry in european phase

Ref document number: 06713408

Country of ref document: EP

Kind code of ref document: A1

WWW Wipo information: withdrawn in national office

Ref document number: 6713408

Country of ref document: EP

NENP Non-entry into the national phase

Ref country code: JP

WWW Wipo information: withdrawn in national office

Country of ref document: JP