The TAG stop codon was replaced with the P2A-iCre cassette. Cre recombinase is expressed under the regulatory control of Calb1 gene elements.

The TAG stop codon was replaced with the T2A-dgCre (DHFR-EGFP-Cre) cassette. Cre recombinase and enhanced green fluorescent protein (EGFP) are expressed under the regulatory control of Calb1 gene elements. The DHFR (dihydrofolate reductase) fragment, derived from the N-terminal 159 amino acids of E. coli ecDHFR, functions as a destabilizing domain (DD). In the absence of specific ligands like TMP (Trimethoprim), this fragment leads to the rapid proteasomal degradation of Cre recombinase. This mechanism allows for the temporal-specific induction of Cre recombinase activity by regulating DHFR-Cre stability with small molecules such as TMP. In addition to studies of Cre recombinase-mediated gene recombination, this model can also be used for EGFP fluorescent protein tracing studies.

Calb1 is located on chromosome 4 of mice. Nuclease Technology was used to design sgRNA; Calb1 knockout mice were obtained by applying high-throughput electroporation of fertilized eggs. After sexual maturity, sperm were collected for cryopreservation.

Zfand4 is located on chromosome 6 of mice. Nuclease Technology will be used to design sgRNA; Zfand4 knockout mice will be obtained by applying high-throughput electroporation of fertilized eggs. After sexual maturity, sperm were collected for cryopreservation.

Otor is located on chromosome 2 of mice. SgRNA and ssDNA will be designed using Nuclease Technology; Otor conditional knockout mice will be obtained by high-throughput electroporation of fertilized eggs. After sexual maturity, sperm will be collected for cryopreservation.

Calb1 is located on chromosome 4 of mice. SgRNA and ssDNA were designed using Nuclease Technology; Calb1 conditional knockout mice were obtained by high-throughput electroporation of fertilized eggs. After sexual maturity, sperm were collected for cryopreservation.