The Wnt signalling pathway plays an important role in regulation of haematopoietic stem cells (HSCs) self-renewal. Purified Wnt3a could remarkably enhance expansion of HSCs. A transgenic bone marrow stromal cells were established by using adenoviral vector mediated Wnt3a gene transfection. The effect of transgenic stromal cells as feeder layer on expansion of CD34+ cells was studied. The group of Wnt3a transfected BM stromal cells with cytokines displays the best expansion effect on of human umbilical cord blood CD34+ hematopoietic stem/progenitor cells in four groups. In vitro, expansion of CD34+ cells, when cocultured with Wnt3a modified stromal cells in the presence of cytokines, was significantly enhanced: CFC by (1.55±0.06) fold; CFC(mix) by (1.95±0.26) fold; HPP-CFC by (1.45±0.40) fold; LTC-IC by (3.83±0.86) fold ,compared with nontransgenic stromal cells with cytokines group. These results strongly suggest that Wnt3a modified BM stromal cells may be a suitable feeder layer for expansion of haematopoietic stem/progenitor cells in vitro.