Objective To construct the eukaryotic expression vector of human HYAL1 gene and obtain MCF-7 and ZR-75-30 cell clones expressing HYAL1 gene stably.   Methods The cDNA encoding HYAL1 gene of human breast cancer was amplified by RT-PCR from the total RNA isolated from human MDA-MB-435S cells and inserted into pcDNA3.1/V5-His-TOPO vector. The recombinant plasmid was transferred into MCF-7 and ZR-75-30 cells.    Results A 1332-bp DNA fragment was successfully amplified from human MDA-MB-435S cell. Restriction enzyme digestion analysis and DNA sequencing showed that HYAL1 gene was inserted into recombinant vector. RT-PCR analysis revealed that HYAL1 gene could be expressed stably in the transfected MCF-7 and ZR-75-30 and it had strong invasive potential.    Conclusion The eukaryotic expression vector of human HYAL1 gene was successfully constructed. MCF-7 and ZR-75-30 cell clones that can express HYAL1 gene were obtained and can promote the invasion.