Dkk3系统性表达转基因小鼠的建立

目的建立系统性表达Dkk3转基因模型小鼠,为研究Dkk3生理功能及对骨生长发育的作用提供工具动物。方法通过ISH来观察Dkk3于C57BL/6J小鼠全身组织中的表达。把Dkk3基因插入系统性表达CMV启动子下游,构建转基因表达载体,显微注射法建立C57BL/6J Dkk3转基因小鼠。PCR鉴定转基因小鼠的基因型,RT-PCR检测Dkk3在骨髓中的表达,Western Blot检测Dkk3在肺脏、脑及肝脏中的表达,BrdU标记染色观察转基因小鼠骨生长情况。结果在生理状态下,Dkk3基因广泛表达,在骨、心脏及脑等组织高表达。建立的2个转基因小鼠品系中,转入的Dkk3基因在骨髓、脑、肝脏及肺组织中均有明显表达。BrdU整合率实验显示转基因小鼠长骨骺区细胞增殖明显低于同龄对照小鼠。结论建立了系统性表达Dkk3转基因小鼠,转入的Dkk3基因明显抑制小鼠长骨骨骺区细胞增殖,为Dkk3对骨生长发育的作用研究提供了有价值的工具动物。

Establisment of Systemic Expression Dkk3 Transgenic Mice

Objective To generate the systemic expression Dkk3 transgenic mice, a model for the study of its function and effects on the development of the bone tissues.Methods The expression pattern of Dkk3 in C57BL/6J mice was observed by tissue in situ hybridization. The transgenic vector was constructed by inserting the murine Dkk3 gene into the down stream of CMV promoter. The transgenic mice were created by the method of microinjection. The genotype of transgenic line was identified by PCR and the expression level of the gene was determined by RT-PCR and Western blot. The growth of the cells in the epiphyseal plate was detected by the BrdU incorporation. Results The endogenous Dkk3 was commonly expressed in mouse, and especially expressed in a higher level in the tissues of bone, heart and brain. Two lines of C57BL/6J transgenic mice with high level of Dkk3 were established. The growth of cells in the epiphyseal plate was inhibited in Dkk3 transgenic mice compared with that of wild type. Conclusion The systemic expression Dkk3 transgenic mouse was successfully established and the transgenic Dkk3 inhibited the cell growth of the epiphyseal plate. The transgenic mouse could be a useful animal model for study of Dkk3 on bone.