Establishment Of NK And NKT Cells-specific Cre Recombinase Knock-in Mice

Gene-targeting technology, including gene knock-out and knock-in, is one of the most important tools for investigating gene function in biomedical research. By using gene-targeting technology, genes can be specifically modified by introducing foreign targeting vectors into mouse ESC genome by homologous recombination. Construction of gene-targeting vectors is the initial step for generating gene-targeted animals.Conditional gene knock-out technology is an important experimental tool that based on the cell or tissue-specific Cre recombinase knock-in mice, to achieve the specific cells and specific gene knock-out stages and to study its function. Using Cre-loxP recombination system to achieve tissue-specific gene knock-out can be completely overcome the traditional gene knock-out technology, which can effectively avoid the complete knock out caused embryonic lethality. In this system, the tissue-specific Cre recombinase knock-in or transgenic mice is an important part.So far, there are many immune cells-specific Cre recombinase knock-in or transgenic mice, such as:B Cell; T Cell ; Dendritic Cell; Macrophage; Neutrophilic granulocyte. But, there is no report of NK and NKT cells-specific Cre recombinase knock-in or transgenic mice.Preparation of tissue-specific Cre recombinase knock-in mice gene, the most common method is to use tissue-specific expression of gene promoters to regulate the expression of Cre recombinase. NK1.1 is known in mice of NK and NKT cell-specific markers, therefore NK1.1 is very suitable as the promoter of the tissue-specific promoter for Cre recombinase mediated by NK and NKT cells in Expression.In this paper, we described the construction of a NK1.1-Cre gene targeting vector, in which a Cre gene was inserted into the downstream of mouse endogenous promoter of NK1.1 gene by using"Recombineering(recombination-mediated genetic engineering)"technique which is a powerful method for fast and efficient construction of vectors for subsequent manipulation of the mouse genome or for use in cell culture experiments. We will introduce this targeting vector into mouse ES cells and identify correctly targeted ES clones by using PCR in the following experiment.Then using Microinjection that putting the correctly targeted ES cells into the F2 generation of offspring's blastocysts(E3.5 days) of the C57BL/6J strain and DBA2 strain, and 10-20 blastocysts implantation in pseudopregnant female mice to obtain chimeric mice. Then the higher rates of male chimeric mice and C57BL/6J female chimeric mice were mated to observe the F1 generation of mice. Identified by PCR genotype of F1 generation of brown mice and to determine the construction of NK1.1-Cre knock-in mouse model.This is the first establishment of NK and NKT cells-specific Cre recombinase knock-in mice. The mice can be widely used in the NK, NKT cells for tissue-specific gene targeting studies and related diseases in the role and mechanism. We forecast in the next few years, there will be more immune cells in different Cre recombinase-specific expression of knock-in or transgenic mice will be established to the different immune cells in gene function of certain specific research.