Reconstructed oocyte of deaf model pig, construction method of reconstructed oocyte, deaf model pig, and construction method and application of deaf model pig

A technology for constructing methods and models, applied in the field of molecular biology and genetic engineering, can solve the problem of not being able to completely replicate the deafness phenotype, and achieve the effect of avoiding phenotypic differences

Pending Publication Date: 2019-11-19
NANJING MEDICAL UNIV +1
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Problems solved by technology

Although the hearing structure and function of mice and other rodents and the phenotype of the mutants are similar to those of humans, they are quite different from humans in terms of developmental genetics, and cannot completely replicate the deafness phenotype of humans.

Method used

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  • Reconstructed oocyte of deaf model pig, construction method of reconstructed oocyte, deaf model pig, and construction method and application of deaf model pig
  • Reconstructed oocyte of deaf model pig, construction method of reconstructed oocyte, deaf model pig, and construction method and application of deaf model pig
  • Reconstructed oocyte of deaf model pig, construction method of reconstructed oocyte, deaf model pig, and construction method and application of deaf model pig

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Embodiment Construction

[0035] The invention provides a method for constructing a deaf pig model pig, which introduces a frameshift mutation into the coding sequence of the OSBP fingerprint motif "EQVSHHPP" of the pig OSBPL2 gene and its upstream coding sequence, thereby accurately simulating the expression of the OSBPL2 gene of a human DFNA67 patient. mutant ( figure 1 ), resulting in the deletion of the OSBP fingerprint motif "EQVSHHPP" and protein truncation, to establish a deafness model pig with OSBPL2 functional deficiency. Specifically, the invention includes constructing reconstituted eggs of OSBPL2 gene knockout model pigs, activating reconstituted eggs and obtaining deaf model pigs through surrogate pregnancy. The reagents or instruments used in the present invention are not indicated by the manufacturer, and they can all be purchased commercially.

[0036] In one embodiment, the construction method of the deafness model pig (flow chart is as follows figure 2 shown) may include but not ...

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Abstract

The invention discloses a reconstructed oocyte of a deaf model pig and a construction method of the reconstructed oocyte, and also discloses the deaf model pig and a construction method and application of the deaf model pig. According to the reconstructed oocyte of the deaf model pig, the construction method of the reconstructed oocyte and the construction method of the deaf model pig, by knockingout a pig OSBPL2 gene, the reconstructed oocyte of a deaf mini-pig is obtained, and in combination with a CRISPR/Cas9 gene editing technology, a somatic cell nuclear transfer technology and an embryotransfer technology, high efficiency and feasibility of the construction method for the genetically engineered deaf model pig are proven. Through the construction method of the deaf model pig, genotypes and deafness phenotypes of human OSBPL2 gene mutations can be precisely duplicated, the pig of which an OSBPL2 gene is knocked out has typical characteristics of human autosomal dominant hereditary hearing loss (DFNA67), therefore, a reliable big animal model can be provided for disease research of the human hereditary hearing loss, and the deaf model pig can be applied to inner-ear gene repairing, inner-ear hair cell regeneration, hearing reconstruction and the like.

Description

technical field [0001] The invention belongs to the fields of molecular biology and genetic engineering, and in particular relates to a reconstructed egg of a deafness model pig and a construction method thereof, a deafness model pig, a construction method and an application thereof. Background technique [0002] Deafness is the most common sensory impairment disease. According to the latest statistics from the World Health Organization (WHO), there are about 466 million hearing disabled people in the world, of which nearly 34 million are children. The incidence of newborn deafness is 0.1-0.3%. 50% of deaf patients are related to genetic factors. According to whether it is accompanied by abnormalities, lesions or systemic diseases outside the ear, hereditary deafness is divided into syndromic hearing loss (SHL) and non-synthetic deafness (Non- syndromic HearingLoss, NSHL), NSHL accounts for about 70%. Autosomal dominant deafness (DFNA) and autosomal recessive deafness (DFNB...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/85C12N15/90C12N9/22C12N15/877C12N5/10A01K67/027A61K49/00
CPCA01K67/0273A01K67/0276A01K2217/075A01K2227/108A01K2267/03A61K49/0008C07K14/47C12N9/22C12N15/85C12N15/8778C12N15/907C12N2800/107C12N2810/10
Inventor 曹新姚俊曾华沙杨海元王盈邢光前陈智斌魏钦俊鲁雅洁戴一凡
Owner NANJING MEDICAL UNIV
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