To investigate the effect of tonifying kidney，benefiting marrow and engendering blood method on JAK2/STAT5 signaling pathway in the erythroid differentiation of hematopoietic stem cells. Methods： On the basis of the previous in vivo animal experiment，the hematopoietic stem cells were cultivated with AA rats serum and induced erythroid differentiation in vitro. The cells were divided into the following seven groups： blank control group，normal control group，model group，stanozolol group，benefiting marrow and engendering blood group，warming kidney and engendering blood group，nourishing kidney and engendering blood group. Erythroid cells RNA and protein were extracted in 4 days respectively，and the level of JAK2，STAT5 mRNA and protein in erythroid cells were detected with RT-PCR and Western blot. Results： Compared with the normal control group，the expressing of JAK2，STAT5 mRNA and protein in cells decreased significantly in the model group(P<0.01).Compared with the model group，the expression of JAK2，STAT5 mRNA and protein both significantly increased in treatment groups(P<0.01 or P<0.05)，while the nourishing kidney group was significantly better than the benefiting marrow group and warming kidney group of the expression of JAK2，STAT5mRNA and protein. Conclusion： The method of tonifying kidney，benefiting marrow and engendering blood can increase the expression of JAK2，STAT5 and promote erythroid differentiation and maturation by affecting JAK2/STAT5 signaling pathway and stimulating activity. The nourishing kidney and engendering blood method is better than benefiting marrow and warming kidney method.