Objective To construct and identify the recombinant eukaryotie expression vector containing specific mutant DNA polβ gene derived from prostate cancer. Methods Total RNA was extracted from prostate cancer tissues with specific DNA polβ mutation and re-versely transcripted to cDNA by universal primer. Utilizing primer with adapter design, the full length DNA poll3 gene with specific mutation sites was amplificated by PCR. T-A clone was constructed and the recombinant was screened and identified. Full length DNA polβ gene with specific mutation sites was then subcloned into expression vector pcDNA3.1 and the recombinant was screened and identified. Results P4 (M1-polβ) and P5 (M2-polβ) were selected. Eukaryotie expression vector peDNA3.1 containing specific mutant DNA polβgene was con-structed and reeombined and two positive colony were acquired by PCR. Conclusions Two eukaryotic expression vectors containing specific mutant DNA poll3 derived with prostate cancer (peDNA3. 1-M1 and peDNA3, 1-M2) are successfully constructed.