Objective:To construct a luciferase reporter plasmid containing the splicing isoform of CD2-associated protein(CD2AP) human gene promoter and its deletants and to evaluate promoter activity in human embryonic kidney (HEK)-293 cells and HeLa cells. Methods:Extract total RNA from stable HeLa cells and make it to be cDNA by RT-PCR. The 2 200 bp fragment of CD2AP002(one of the spliceosome of CD2AP)was amplified by PCR with cDNA as a template and was directionally cloned into pGL3-Basic multiple cloning sites to construct a new luciferase reporter plasmid. We repeat the above steps to build a series of 5′ deletion promoter plasmids. Transfection of HEK 293 cells and HeLa cells with these new plasmids was performed to induce the relative luciferase activity. Results:DNA sequencing and restriction endonuelease analysis verified the successful construction of the plasmid contaning CD2AP002 human gene promoter and its deletants. Conclusion:The plasmid pGL3-CD2AP002 promoter and its deletants are successfully constructed and have some promoter activities in HEK293 cells and HeLa cells.