Conditional Knockout and Recombinant Cre/loxP, Flp/FRT System
To do this experiment, I have checked some conditional gene knockout, Cre / loxP and FLP / frt recombination system materials and make a summary.
Classical gene knockout is sometimes unable to give birth due to the effects of the gene on the development of the embryo, or the experimental animals die of severe physiological defects after birth, or the animals can not produce offspring to obtain a homozygous animal model. These can be avoided through conditional gene knock-out or knock in. That is to say, in specific cells or tissues and specific cell development stages a particular gene is knocked in. It has important significance to study the features of specific gene in specific organization.
Currently Cre/loxP and Flp/FRT restructuring system is wildly used.
Cre / LoxP and Flp - LRT system derived from yeast can be used to study the phenotype of target gene inactivation in specific tissues and organs. Through the conventional gene targeting, equipping two loxP aligned in the same direction in the genomic target sites and mounting loxP floxed produced by ES cells. Then making the loxP floxed mice and CRE transgenic mice hybridize to produce modified conditional mutant mice. In the "floxed loxP" mice, although the target gene has been loaded on both sides of a loxP, but the target gene has not changed. So the "floxed 1oxP" mouse phenotype is still the same as the wild type’s. But when it crosses with Cre transgenic mice, the offspring will have "floxed loxP" target gene and Cre gene. The Cre gene expresses to produce Cre recombinant which can mediate the 1oxP of the target gene, cut a loxP and a target gene. Thus, the modification of the target gene is based on the expression of Cre. The expression characteristic of Cre determines the modification of target gene. That means which cells the Cre choose to express, where the modification of target gene will happen. The expression level of Cre will affect the efficiency of target gene in the cells. So as long as the expression specificity and the expression level can be controlled, it is possible to achieve the specificity and extent of target gene modification in mice.
Reference: knock in mouse