Objective：To construct eukaryotic expression vectｏr carrying anti-sense PRLR gene and investigate whether the reco-mbined plasmids could regulate the expression of PRLR mRNA and affect the growth of human breast cancer cell line MCF-7. Methodｓ：PRLR cDNA was inserted into the expression plasmid pcDNA3.1（-） to construct the recombined plasmids p-anti-PRLR that encoding PRLR cDNA in an anti-sense orientation, then the recombined plasmids transfected MCF-7 cells by lipofectin. The stable cell lines were obtained after G418 selection. PRLR mRNA expression was detected by real-time PCR. MTT assay, clone formation test, and the flow cytometry（FCM） were used to evaluate cell proliferation. Resultｓ：（1）The eukaryotic expression vector p-anti-PRLR was constructed successfully. （2）Antisense PRLR gene decreased the level of PRLR mRNA in MCF-7 cells. （3）After antisense gene transfection, the proliferation ability of MCF-7 cell was decreased, and the G1 phase cells were increased, but the S phase cells were decreased in cell cycle. Conclusion：The results suggested that antisense PRLR could inhibit MCF-7 cell proliferation, and it might be significant for the breast cancer gene therapy.