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Abstract  Transgenic GFP gene mice are widely used. Given the unique advantages of immunodeficient animals in the field of oncology research, we aim to establish a nude mouse inbred strain that stably expresses enhanced GFP (EGFP) for use in transplanted tumor microenvironment (TME) research. Female C57BL/6‐Tg(CAG‐EGFP) mice were backcrossed with male BALB/c nude mice for 11 generations. The genotype and phenotype of novel inbred strain Foxn1 nu .B6‐Tg(CAG‐EGFP) were identified by biochemical loci detection, skin transplantation and flow cytometry. PCR and fluorescence spectrophotometry were performed to evaluate the relative expression of EGFP in different parts of the brain. Red fluorescence protein (RFP) gene was stably transfected into human glioma stem cells (GSC), SU3, which were then transplanted intracerebrally or ectopically into Foxn1 nu .B6‐Tg(CAG‐EGFP) mice. Cell co–expression of EGFP and RFP in transplanted tissues was further analyzed with the Live Cell Imaging System (Cell’R, Olympus) and FISH. The inbred strain Foxn1 nu .B6‐Tg(CAG‐EGFP) shows different levels of EGFP expression in brain tissue. The hematological and immune cells of the inbred strain mice were close to those of nude mice. EGFP was stably expressed in multiple sites of Foxn1 nu .B6‐Tg(CAG‐EGFP) mice, including brain tissue. With the dual‐fluorescence tracing transplanted tumor model, we found that SU3 induced host cell malignant transformation in TME, and tumor/host cell fusion. In conclusion, EGFP is differentially and widely expressed in brain tissue of Foxn1 nu .B6‐Tg(CAG‐EGFP), which is an ideal model for TME investigation. With Foxn1 nu .B6‐Tg(CAG‐EGFP) mice, our research demonstrated that host cell malignant transformation and tumor/host cell fusion play an important role in tumor progression.

Novel enhanced GFP‐positive congenic inbred strain establishment and application of tumor‐bearing nude mouse model