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目的:克隆小鼠CXCR4基因启动子,并建立CX-CR4报告基因系统。方法:设计合成PCR引物,从小鼠基因组DNA中扩增并克隆小鼠CXCR4基因的启动子区。序列测定确认后,用克隆的启动子片段构建CXCR4荧光素酶报告基因载体pGL-CXCR4。转染细胞,用双报告基因系统检测报告基因载体的活性。结果:成功扩增了小鼠CXCR4基因的启动子区。克隆入质粒载体后经DNA序列测定证实了其序列。通过转染细胞和荧光素酶分析,证实所构建的报告基因可以反映CXCR4启动子的活性。结论:成功扩增、克隆了小鼠CXCR4基因启动子,并成功建立起其报告基因系统,为后续的研究奠定了基础。
OBJECTIVE: To clone mouse CXCR4 gene promoter and establish CX-CR4 reporter system. Methods: PCR primers were designed and synthesized. The promoter region of mouse CXCR4 gene was amplified and cloned from mouse genomic DNA. After sequencing confirmation, the CXCR4 luciferase reporter gene vector pGL-CXCR4 was constructed using the cloned promoter fragment. The cells were transfected and the dual reporter system was used to test the activity of the reporter vector. Results: The promoter region of mouse CXCR4 gene was successfully amplified. After cloning into plasmid vector, its sequence was confirmed by DNA sequencing. The transfected cells and luciferase analysis confirmed that the constructed reporter gene can reflect the activity of CXCR4 promoter. Conclusion: The mouse CXCR4 gene promoter was successfully amplified and cloned, and its reporter gene system was successfully established, which laid the foundation for the follow-up study.