Method for preparing gene knockout dog by adopting somatic cell cloning technology

Abstract

The invention relates to a method for preparing a gene knockout dog by adopting a somatic cell cloning technology, and particularly relates to a method for preparing a gene knockout dog by adopting alow osmotic pressure fusion solution and an autologous embryo transplantation technology.

Description

technical field [0001] The present invention relates to a method for preparing a gene-knockout dog by using somatic cell cloning technology, in particular to a method for preparing a gene-edited dog by using the somatic cell cloning technology of low-osmotic fusion fluid and autologous embryo transfer. Background technique [0002] Dogs are currently one of the most commonly used experimental animals in basic medical research and teaching, especially playing an important role in experimental research in physiology, pharmacology and pathophysiology. Through the whole genome sequencing analysis of dogs, a total of about 19,300 genes have been identified, of which about 18,000 genes are the same as known human genes, and the similarity between the genome and human is higher than that of other experimental animals such as mice. Dogs are also very similar to humans in terms of genetic diseases. There are more than 360 genetic diseases such as cancer, heart disease, deafness, blin...

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Problems solved by technology

[0005] However, because the reproductive physiology of dogs is very different from that of other mammals, it is extremely difficult to operate canine oocytes and embryos in vitro. The establishment of gene knockout or transgenic modified model dogs and somatic cell cloned dogs is usually considered a Difficult to succeed

[0006] In summary, in the existing techniques for preparing gene knockout dogs: (1) usually use canine normal fertilized embryos for cytoplasmic injection. During the splitting process, the gene knockout types of different blastomeres are likely to be inconsistent, that is, it is possible to obtain chimeras of gene knockout dogs; (2) gene knockout dogs have not been obtained by gene targeting technology, and gene knockout dogs have not been tested. Cell cloning to obtain a stable gene-knockout cloned dog with a gene knockout efficiency of 100%; and (3) female dogs with synchronous estrus are often selected for allogeneic embryo transfer, the success rate is low, resulting in an increase in the production cost of cloned dogs, and , the number of female dogs in estrus required for allogeneic embryo transfer is large, and the operation methods are complicated, so it is difficult to achieve the goal of actual industrial production of cloned dogs

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