AIM To study the hematopoietic reconstitution ability of fresh and cultured cord blood CD34(+) cells by sublethally irradiated NOD/SCID mice. METHODS Mononuclear cells (MNC) were separated from cord blood and then cultured with stem cell factor (SCF)+ flt3 ligand (FL)+ thrombopoietin (TPO)+ interleukin-3 (IL-3)+ IL-6 for 14 d. CD34(+) cells were isolated from fresh or cultured MNC using miniMACS magnetic separation system. Then 4x10(5) CD34(+) cells together with 5x10(6) CD34(-) cells were injected into NOD/SCID mice. After transplantation, the dynamics of hematopoieitc recovery in the mice were measured. After 6 weeks, bone marrow (BM) and spleen samples were obtained from mice for detecting human cells. RESULTS The number of total cells increased 1.78-fold after 14 d of culture. All recipients received fresh and cultured CD34(+) cells survived. Human cells, human lineage cells and human ALU sequence could be detected in BM and spleen. The peripheral blood counts of all transplanted mice recovered. The engraftment levels of cultured CD34(+) cells were similar to those of fresh CD34(+)cells but the percentage of human lineage cells was higher than that of fresh CD34(+) cells. CONCLUSION CD34(+) cells cultured for 14 d still preserved the hematopoietic reconstitution ability and showed better multilineage reconstitution ability than fresh CD34(+) cells.