Abstract: Objective To construct mouse granulocyte-macrophage colony-stimulating factor (mGM-CSF) expression vector and express, purify and refold mGM-CSF protein. Method Based on previously constructed fusion protein hIL-2/mGM-CSF expression vector, pET-11c/mGM-CSF expression vector was constructed routinely and transformed into BL21 (DE3). The inclusion body protein was washed with our patented method and refolded with renaturation buffer containing low-concentration guanidinium chloride (Gu·Cl). The refolded protein was purified with affinity chromatography. Results pET-11c/mGM-CSF vector was constructed successfully. The host bacteria was cultured in TH broth and induced with 0.1 mmol/L IPTG at 32 ℃, which resulted in the expression level of 60.6%. The best refolding effect was achieved with the renaturation media containing glutathione and 1.5 mol/L Gu·HCl. After purification with affinity chromatography, the purity of the target mGM-CSF protein reached 95% with activity of 5×106 U/mg. Conclusion Engineered bacteria BL21/pET- 11c/mGM-CSF with efficient mGM-CSF expression and laboratory scale renaturation and purification of mGM-CSF have been established, which facilitates further researches into the anti-tumor function of the dendritic cells and GM-CSF in vivo.