Method for constructing severe immunodeficiency and liver injury dual pig model and application

Abstract

The invention relates to a method for constructing a severe immunodeficiency and liver injury dual pig model and application, and belongs to the technical field of animal biology. According to the method, RAG2, IL2RG and FAH genes in porcine fetal fibroblasts are knocked out by adopting a CRISPR/Cas9 technology, RAG2-/-/IL2RG-/Y/FAH-/-three-gene editing cloned pigs are constructed by utilizing a somatic cell nuclear transplantation technology, and the severe immunodeficiency and liver injury dual pig model is obtained through phenotype analysis and identification. The problems of long production cycle, low efficiency, irreversible injury, lack of immunodeficiency and non-ideal humanization degree of simultaneous application of liver injury and the like in the existing model construction technology are overcome, and batch construction of the severe immunodeficiency and liver injury dual pig model can be realized through a continuous cloning technology, and the method has huge advantages and potential market application prospects in the related fields of tumor biology, cell transplantation, humanized animal models and the like.

Description

technical field [0001] The invention belongs to the field of animal biotechnology, and in particular relates to a method and application for constructing a double pig model of severe immunodeficiency and liver injury. Background technique [0002] Immunodeficient animals play an important role in the fields of biomedical and basic medical research. Liver transplantation is an important treatment for end-stage liver failure, and the shortage of liver donors is the main bottleneck restricting the clinical application of liver transplantation. As one of the important ways to solve the serious shortage of liver organs, hepatocyte transplantation technology can produce humanized liver by implanting human hepatocytes in animal models of immunodeficiency and liver injury, and obtaining humanized hepatocytes in batches can solve the problem. The limited source of primary hepatocytes has played an important role in the treatment of human hepatitis B (HBV) and hepatitis C (HCV) disea...

AI Technical Summary

Problems solved by technology

[0004] (1) Rodents such as mice are distantly related to humans, have large body size differences, and have short lifespans. It is difficult to simulate human physiological and metabolic processes with rodents such as mice. The application in the field of scientific research has been greatly restricted

[0005] (2) At present, the construction of dual mouse models uses more traditional gene editing methods, and even chemical drugs are used in the preparation of liver injury, which has irreversible damage, and the success rate of modeling is low, which is not conducive to subsequent simulations Human Surgical Procedures and Preclinical Evaluation

[0006] (3) FAH obtained by hybridization methods commonly used - / - / Rag - / - / IL2RG - / - A mouse model of simultaneous three-gene modification, first by Rag - / - / IL2RG - / - based on the FAH - / - Mice mated to produce Rag - / + / IL2RG - / + / FAH - / + individuals, and then obtain FAH by selfing - / - / Rag - / - / IL2RG - / - Individuals modified by the three genes at the same time, and some are even chimeric mice obtained by injection of fertilized eggs. This method takes a long time to construct and the production efficiency of the model is low.

[0007] (4) Current mouse FAH - / - / Rag - / - / IL2RG - / - In the model, there is no way to remove NK cells, and the inflammatory response is different from that of humans, so it cannot reshape the human model very well.

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