H11-CAG-MerCreMer Mouse

H11-CAG-MerCreMer mice were crossed with Rosa26-LSL-tdTomato mice to generate heterozygous offspring expressing both MerCreMer recombinase and the LSL-tdTomato fluorescent reporter. Following tamoxifen induction, Cre recombinase efficiently excises the stop element (LSL) upstream of the tdTomato sequence, resulting in the expression of the tdTomato fluorescent protein in Cre-positive cells. Peripheral blood was collected 7 days post-induction for flow cytometry (FACS) analysis to assess tdTomato expression. Additionally, tissues from the lung, liver, brain, spleen, kidney, stomach, heart, and rectum were harvested for fluorescence microscopy, enabling the evaluation of Cre recombinase activity across various tissues.

Cre+Tam+：CAG-MerCreMer[KI/+];Rosa26-LSL-tdTomator[CKI/+]；Tamoxifen;

Cre+Tam-：CAG-MerCreMer[KI/+];Rosa26-LSL-tdTomator[CKI/+]；Corn oil;

Cre-Tam+：Rosa26-LSL-tdTomato[CKI/CKI]；Tamoxifen.

①: Robust tdTomato expression was detected in peripheral blood CD45+CD11b+ cells, with a recombination efficiency of 99.68%. In CD45+CD3+ T cells, the recombination efficiency was 85.54%; in CD45+CD19+ B cells, it was 75.13%; and in CD3-CD335+ NK cells, the efficiency reached approximately 96.16%.

②: Recombination efficiency in CD45+CD11b+ cells was 98.95%, while in T cells it was 63.38%, in B cells 66.76%, and in NK cells approximately 91.05%.

③: CD45+CD11b+ cells showed a recombination efficiency of 99.6%. In T cells, the efficiency was 76.61%, in B cells 89.89%, and in NK cells around 89.52%.

④: Recombination efficiency in CD45+CD11b+ cells was 98.79%, while in T cells it was 64.05%, in B cells 90.22%, and in NK cells approximately 86.27%.

⑤~⑥: In the corn oil-treated control group (Cre+Tam-), minimal tdTomato expression leakage was observed in peripheral blood immune cells, whereas in the Cre-Tam+ group, leakage was nearly undetectable.

In Cre+Tam+ mice, strong orange autofluorescence was observed in the alveoli, bronchial epithelial cells, smooth muscle fibers, and liver tissues, indicating effective Cre-mediated recombination in these cell types. In the control group (Cre+Tam-), minimal expression leakage was detected in the lung, with some leakage observed in the liver.

In Cre+Tam+ mice, partial orange autofluorescence was detected in the third ventricle, fourth ventricle, olfactory bulb, and cerebellum of the brain. In the spleen, robust orange autofluorescence was seen in the white pulp, red pulp, and trabeculae, reflecting Cre recombination in these tissues.

In Cre+Tam+ mice, significant orange autofluorescence was observed in the tubular epithelial cells and glomeruli of the kidneys, as well as in the mucosal and muscular layers of the stomach, indicating successful recombination in these tissues. Minimal expression leakage was noted in the stomach tissues of Cre+Tam- control mice.

In Cre+Tam+ mice, significant orange autofluorescence was detected in the heart tissues, with autofluorescence signals also evident in the rectum, indicating Cre recombination in these regions.

In H11-CAG-MerCreMer mice treated with tamoxifen, substantial Cre recombinase activity was observed in peripheral blood CD45+CD11b+, CD45+CD3+, CD45+CD19+, and CD45+CD335+ cells. Strong fluorescent signals indicating successful recombination were also detected in the lungs, liver, spleen, kidneys, stomach, and heart, with partial expression seen in the brain and rectum. Based on these findings, we recommend a tamoxifen induction protocol of 12.5 mg/kg at 24-hour intervals for 5 doses, followed by tissue collection 7 days post-induction to achieve optimal results. This model may have an elevated sensitivity to tamoxifen induction. We recommend customizing the induction protocol and conducting pilot studies to minimize risks. If the target gene impacts survival or phenotype, consult relevant literature and tailor induction protocols accordingly. Validation results are for reference and may vary across studies.