Hairless model pig reconstructed oocyte and reconstruction method thereof and model pig construction method

Abstract

The invention relates to mutation site controllable hairless model pig reconstructed oocyte and a reconstruction method thereof and a model pig construction method. According to the hairless model pig reconstructed oocyte and reconstruction method thereof and the model pig construction method, by utilizing CRISPR/Cas9 and point mutation technology, cells can be accurately repaired by utilizing provided ssODNs as a template in a homologous recombination manner after Cas9 is cut at the gene specific site, and a termination codon is introduced at the gene specific site through the CRISPR/Cas9 and ssODNs design, so that gene translation is early terminated, and further the loss of gene function can be caused. Human disease gene mutation type can be accurately replicated by utilizing the method, so that the phenotype difference caused due to different gene mutation forms can be avoided. A hairless model pig constructed by utilizing the method has no other abnormities except for hair loss, hoof development defect and filiform papillae development defect, and can be used for various dermatological researches and external skin drugs and cosmetics. The skin transformed immunologically can be used for clinical skin transplantation.

Description

technical field [0001] The invention relates to the field of animal model construction, in particular to a reconstituted egg of a hairless model pig, a construction method thereof, and a construction method of a model pig. Background technique [0002] Biomedical research cannot do without animal models. Unlike humans, other mammals are mostly covered with thick hair, which is not conducive to dermatological research. Establishing a hairless animal model, in addition to avoiding skin damage caused by hair removal during the experiment, is mainly used for skin transplantation, skin aging and anti-aging research, cosmetics and external skin drug testing. In addition, hairless animals can be used as research models for alopecia, low-onset diseases. According to investigations, the incidence rate of male hair loss in my country is as high as 25%. With the acceleration of the pace of life, the enhancement of work pressure, and the change of diet and living habits, the incidenc...

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Problems solved by technology

[0006] Before the emergence of high-efficiency gene targeting technologies such as TALEN and CRISPR / Cas 9, the efficiency of site-specific gene modification in large animals was extremely low, because there was a lack of embryonic stem cell (ES) lines in large animals such as pigs, and only somatic cells could be gene-targeted after gene targeting. Somatic cell nuclear transfer is used to produce large animals for gene targeting. Since the efficiency of somatic cell homologous recombination is far lower than that of embryonic stem cells, it is very difficult to target somatic cells.

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