LU600477B1 - An sgrna targeting the knockout of the zebrafish sigirr gene and its crispr/cas9 system and applications - Google Patents

An sgrna targeting the knockout of the zebrafish sigirr gene and its crispr/cas9 system and applications

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Publication number
LU600477B1
LU600477B1 LU600477A LU600477A LU600477B1 LU 600477 B1 LU600477 B1 LU 600477B1 LU 600477 A LU600477 A LU 600477A LU 600477 A LU600477 A LU 600477A LU 600477 B1 LU600477 B1 LU 600477B1
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LU
Luxembourg
Prior art keywords
zebrafish
sigirr
knockout
gene
crispr
Prior art date
Application number
LU600477A
Other languages
English (en)
Inventor
Huizhi Guo
Kai Luo
Xincheng Jiang
Qiaoqing Xu
Simei Hu
Original Assignee
Univ Yangtze
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 Univ Yangtze filed Critical Univ Yangtze
Application granted granted Critical
Publication of LU600477B1 publication Critical patent/LU600477B1/en

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    • 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
    • A01K67/0276—Knock-out vertebrates
    • 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
    • C12N15/11—DNA or RNA fragments; Modified forms thereof; Non-coding nucleic acids having a biological activity
    • C12N15/113—Non-coding nucleic acids modulating the expression of genes, e.g. antisense oligonucleotides; Antisense DNA or RNA; Triplex- forming oligonucleotides; Catalytic nucleic acids, e.g. ribozymes; Nucleic acids used in co-suppression or gene silencing
    • C12N15/1138—Non-coding nucleic acids modulating the expression of genes, e.g. antisense oligonucleotides; Antisense DNA or RNA; Triplex- forming oligonucleotides; Catalytic nucleic acids, e.g. ribozymes; Nucleic acids used in co-suppression or gene silencing against receptors or cell surface proteins
    • 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
    • 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
    • C12N9/00—Enzymes; Proenzymes; Compositions thereof; Processes for preparing, activating, inhibiting, separating or purifying enzymes
    • C12N9/14—Hydrolases (3)
    • C12N9/16—Hydrolases (3) acting on ester bonds (3.1)
    • C12N9/22—Ribonucleases [RNase]; Deoxyribonucleases [DNase]
    • 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
    • A01K2217/07—Animals genetically altered by homologous recombination
    • A01K2217/075—Animals genetically altered by homologous recombination inducing loss of function, i.e. knock out
    • 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
    • A01K2227/40—Fish
    • 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
    • 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
    • A01K2267/0337—Animal models for infectious diseases
    • 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
    • C12N2310/00—Structure or type of the nucleic acid
    • C12N2310/10—Type of nucleic acid
    • C12N2310/20—Type of nucleic acid involving clustered regularly interspaced short palindromic repeats [CRISPR]
    • 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
    • C12N2800/10—Plasmid DNA
    • C12N2800/106—Plasmid DNA for vertebrates
    • C12N2800/107—Plasmid DNA for vertebrates for mammalian

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

Claims (10)

REVENDICATIONS LU600477
1. Un sgRNA ciblant l'inactivation du gène SIGIRR du poisson zèbre, caractérisé en ce que la séquence nucléotidique du sgRNA est représentée dans SEQ ID NO : 1.
2. Un ensemble d'amorces pour amplifier le sgRNA selon la revendication 1, caractérisé en ce que l'ensemble d'amorces comprend une amorce en amont représentée dans SEQ ID NO : 2 et une amorce en aval représentée dans SEQ ID NO:
3.
3. Un système CRISPR/Cas9 ciblant l'inactivation du gène SIGIRR du poisson zèbre, caractérisé en ce que le système CRISPR/Cas9 comprend le sgRNA selon la revendication 1 et l'ARNm Cas9.
4. Le système CRISPR/Cas9 selon la revendication 3, caractérisé en ce que le procédé de préparation de l'ARNm Cas9 comprend les étapes suivantes : linéarisation du plasmide pXT7-zCas avec Xba |I-QC, récupération du produit de digestion et transcription in vitro pour synthétiser l'ARNm Caso.
5. Kit de ciblage de l'inactivation du gène SIGIRR du poisson zèbre, caractérisé en ce qu'il comprend le système CRISPR/Cas9 selon la revendication 3 ou la revendication
4.
6. Procédé d'inactivation du gène et de reproduction de poissons zèbres inactivés du gène SIGIRR, caractérisé en ce qu'il comprend les étapes suivantes : introduction du système CRISPR/Cas9 selon la revendication 3 ou la revendication 4 ou du système CRISPR/Cas9 selon la revendication 5 dans des embryons de poisson zèbre, culture et sélection pour obtenir des poissons zèbres inactivés du gène SIGIRR.
7. Procédé selon la revendication 6, caractérisé en ce qu'il comprend en outre 44600477 étapes suivantes : les poissons zèbres inactivés du gène SIGIRR sont hybridés avec des poissons zèbres de type sauvage pour obtenir des embryons de génération F1, qui sont cultivés et sélectionnés pour obtenir les poissons zèbres de génération F1 inactivés du gène SIGIRR ; les poissons zèbres de génération F1 inactivés du gène SIGIRR sont croisés pour obtenir des poissons zèbres homozygotes inactivés du gène SIGIRR.
8. Application du poisson zèbre avec gène SIGIRR knockout obtenu par le procédé de la revendication 6 ou de la revendication 7 à l'étude des maladies infectieuses bactériennes chez les poissons.
9. Application du poisson zèbre avec gène SIGIRR knockout obtenu par le procédé de la revendication 6 ou de la revendication 7 au criblage de médicaments liés à une infection antibactérienne chez les poissons.
10. Application de la revendication 8 ou de la revendication 9, caractérisée en ce que les bactéries sont Edwardsiella piscicida ou Mycobacterium marinum.
LU600477A 2024-03-07 2025-03-06 An sgrna targeting the knockout of the zebrafish sigirr gene and its crispr/cas9 system and applications LU600477B1 (en)

Applications Claiming Priority (1)

Application Number Priority Date Filing Date Title
CN202410263027.3A CN118147139A (zh) 2024-03-07 2024-03-07 一种靶向敲除斑马鱼SIGIRR基因的sgRNA及其CRISPR/Cas9系统和应用

Publications (1)

Publication Number Publication Date
LU600477B1 true LU600477B1 (en) 2025-09-10

Family

ID=91289972

Family Applications (1)

Application Number Title Priority Date Filing Date
LU600477A LU600477B1 (en) 2024-03-07 2025-03-06 An sgrna targeting the knockout of the zebrafish sigirr gene and its crispr/cas9 system and applications

Country Status (2)

Country Link
CN (1) CN118147139A (fr)
LU (1) LU600477B1 (fr)

Also Published As

Publication number Publication date
CN118147139A (zh) 2024-06-07

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