Bone is one of the most common sites of metastasis in patients with advanced breast cancer; however, the mechanisms of bone metastasis remain to be fully elucidated. Animal models are essential research tools for investigating the mechanisms of diseases and drug actions. To date, there have only been a few reports in which C57BL/6 mice were used for the study of bone metastases of breast cancer. In the current study, we found that intracardiac inoculation of C57BL/6 mouse-derived parental E0771 breast cancer cells (E0771/Pa) frequently lead to bone metastases in C57BL/6 mice within 2 weeks. The bone-metastatic clone of E0771 (E0771/Bone) established by sequential in vivo selection demonstrated a higher bone-metastatic potential. Although there were no apparent differences in cell morphology or proliferation in monolayer cultures, E0771/Bone showed increased tumorsphere formation in suspension cultures and tumor formation in the orthotopic mammary fat pad in C57BL/6 mice compared with E0771/Pa. Furthermore, E0771/Bone expressed breast cancer stem-like cell surface markers CD24-/CD44+. These findings suggest that E0771/Bone possesses cancer stem-like properties. Quantitative PCR analysis revealed that mRNA expression of parathyroid hormone-related protein (PTHrP), the most common mediator of osteolytic bone metastases of breast cancer, was significantly upregulated in E0771/Bone. Thus, cancer stem-like properties and elevated PTHrP expression likely contribute to the enhanced metastatic potential of E0771/Bone. We believe that this new mouse model is a useful tool for in vivo studies of bone metastases of breast cancer, especially for those using genetically engineered mice with a C57BL/6 background.