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Conditional point mutation mouse model-induced

1.Definition

A mouse model where a point mutation is introduced into the target gene of specific cells or tissues at a specific time by effect of the inducer. Before the induction, the wildtype protein is expressed; after the induction, the mutated protein is expressed in all of the body's specified cells or tissues.


2.Advantages and disadvantages

Advantages: this strategy can avoid a deadly phenotype caused by a conventional point mutation, such as death of the embryonic stem cell, death of the chimeric mouse, or death of the generation F1 heterozygous mouse.

The model mouse grows and develops normally, and the mouse expresses the mutated protein when given the inducer, thereby allowing timed control of the protein's expression.

Disadvantages: ensure the mutated gene's expression level and expression profile is accordant with the wildtype gene's expression when designing.


3. Principle

The induced point mutation system is based on a receptor that translocates from the cytoplasm to the cell nucleus to regulate gene expression after the cell nucleus hormone receptor binds to the corresponding ligand. The estrogen receptor (ER) is this type of cell nucleus hormone receptor, which can bind to the estrogen and the antagonist of the estrogen-Nolvadex. When inactivated, the estrogen receptor will bind to the heat shock protein 90 (Hsp90) in the cytoplasm, and cannot enter the cell nucleus. When estrogen or Nolvadex binds to the estrogen receptor, the ER is separated from Hsp90 and can enter the cell nucleus. The intracellular localization of the Cre recombinase that can only bind to loxP locus in cell nucleus can be controlled with this ER. The Cre-ER fusion protein without estrogen present remains in the cytoplasm due to the binding of ER and Hsp90. However, when the Nolvadex is added, the Cre-ER fusion protein is separated from Hsp90 and enters the cell nucleus. In the cell nucleus, Cre binds to the floxed (loxP-flanked) target gene to effect recombination, and the gene knockout animal model is obtained. This system uses Nolvadex, a modified estrogen receptor, to avoid the binding of the estrogen receptor with endogenous estrogen, and ensure timing control of Cre. The expression of Cre-ER fusion protein is regulated by a tissue specific promoter.

 

4. Application

Lethal target gene study

Study of target gene in development or age-related diseases

Drug target affirmation

Preclinical drug evaluation


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