An animal model and a method for producing an animal model

A model, non-human animal technology, applied in the field of non-human animal models, can solve problems such as the generation of pig models

Inactive Publication Date: 2009-12-30
AARHUS UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0021] However, since pigs and humans belong to a different evolutionary clade, the introduction of mutations known to cause mutations in human-specific genes for disease cannot be expected to produce the desired phenotype in the pig model

Method used

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  • An animal model and a method for producing an animal model
  • An animal model and a method for producing an animal model
  • An animal model and a method for producing an animal model

Examples

Experimental program
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Embodiment 1

[0341] A Transgenic Pig Model of Amyotrophic Lateral Sclerosis (ALS)

[0342] To create a transgenic pig that can be used as a potential pig model for the human neurodegenerative disease ALS, the mutation Gly93Arg was introduced into the gene of pig SOD1 by site-directed mutagenesis. Mutations were chosen based on protein structure inference, as the crystal structure of human SOD1 revealed an extremely condensed structure, which suggested that substitutions with large charged arginines at the positions of small, relatively flexible glycines could potentially cause protein Major changes inside. Furthermore, several different substitutions at this position cause ALS in humans, including the G93R mutation [45, 46]. To prevent the possibility of truncating important elements within the DNA construct after the SMGT step, the DNA fragment already containing the porcine SOD1 cDNA contained additional 5'- and 3'-guide nucleotides of the CMV promoter and SVpolyA fragment. In general,...

Embodiment 2

[0453] Transgenic pig model for Parkinson's disease

[0454] Cloning of porcine SNCA cDNA

[0455] Full-length porcine α-synucleus (SNCA) cDNA was isolated from the cerebellum by a combination of RT-PCR and RACE. Initially, blast searches were performed using the human SNCA cDNA sequence using GenBank (http: / / www.ncbi.nlm.nih.gov / Genbank / index.html) and the porcine EST database of The Danish Institute of Agricultural Sciences (DIAS). Use NCBI Blastall, select blastn, and the lowest value is 10-8 to search for sequence similarity with gapped alignment. The porcine cDNAs thus identified were used to obtain oligonucleotide primers for the clones and as interrogation conditions for further searches in the local genome DIAS sequence database.

[0456] The porcine cerebellum tissue used for RT-PCR cloning of porcine SNCA was obtained from adult pigs. After resection, the tissue was minced into pieces and pulverized in liquid nitrogen. Total RNA was isolated by the RNeasy method ...

Embodiment 3

[0542] Transgenic Pig Model Animals of Alzheimer's Disease

[0543] PSEN1 and PSEN2 isolation and sequencing

[0544]Pig brain, lymphocyte and liver RNA was isolated using TRI-reagent (Sigma). For RT-PCR of PSEN1, the following primers were used (PSEN1 forward, 5'-TGGAGGAGAACACATGAAAGAAAG-3' (SEQ ID NO:95); PSEN1-forward-EcoR 15'-GGGGAATTCTGGAGGAGAACACATGAAAGAAAG-3' (SEQ ID NO:96 ); PSEN1 reverse EcoR1, 5'-GGGGAATTCCCTGACTTTGTTAGATGTGGACAC-3' (SEQ ID NO: 97). The reverse primer was used to incubate at 50°C for RT-PCR reaction for 60 minutes, and then PSEN1 forward-EcoR1 and PSEN1 reverse -EcoR1 primer for PCR with reaction conditions: 3 min at 94°C, 35 cycles of 45 s at 94°C, 30 s at 62°C, 2 min at 68°C, followed by a final extension step at 68°C for 7 min The amplified DNA fragments were purified by agarose gel and sequenced directly or EcoR1 cloned into pCDNA3, followed by DNA purification and sequencing. For RT-PCR of PSEN2, the following primers were used (PSEN2-forward,...

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Abstract

The present invention discloses a non-human animal model for a hereditary autosomal dominant disease. The non-human animal model expresses at least one phenotype associated with the disease and is obtained by a genetic determinant. The invention also relates to sperm cells and embryos comprising the genetic determinant for an autosomal dominant disease. Furthermore, methods for producing the non-human animal model, sperm cell, and embryos are disclosed.

Description

[0001] All patent and non-patent documents cited in this application are incorporated by reference in their entirety. technical field [0002] The present invention relates to non-human animal models, such as porcine animal models, and methods of producing non-human animal models by sperm-mediated gene transfer (SMGT). This non-human animal model can be used to study genetic disorders involved in genetic disorders, such as autosomal dominant inheritance, when the genetic determinants involved are used in SMGT, i.e., when the genetic determinants confer a dominant phenotype on said genetic disorder Diseases, such as protein conformation disorders, such as amyotrophic lateral sclerosis (ALS), Alzheimer's disease, Parkinson's disease, trinucleotide repeat-associated diseases, Huntington's disease, and achondroplasia. Background of the invention [0003] Transgenic non-human animals can be used to understand the role of individual genes in the whole animal body and the interrela...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/85C12N15/877
CPCC12N15/8778A01K2267/03G01N33/5088A01K2217/00A01K2227/108A01K2267/035C07K14/4711C12N15/8509A61K49/0008A01K2267/0318A01K2267/0312A01K67/0275A01K2267/0306A01K2217/056
Inventor 罗尼·布鲁恩·麦德森克里斯蒂安·本迪克森克努德·拉森康妮·亚科布森朱尔博·汤姆森
Owner AARHUS UNIV
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