The auto-inducing expression of mutant human glucagon-like peptide-1 fusion protein in Escherichia coli BL21 (DE3) was studied. In order to increase the protein yield, we optimized both the auto-induction fermentation conditions and medium. The results showed that the optimal auto-induction medium is: tryptone 19.17 g/L, yeast extract 9.59 g/L, Na2HPO4 5.72 g/L, KH2PO4 5.48 g/L, (NH4)2SO4 2.66 g/L, NaCl 3.33 g/L, glycerol 2% (v/v), glucose 0.68 g/L, lactose 6.33 g/L, MgSO4 0.24 g/L. The genetically engineered strain was cultivated at 33°C, pH7, in a 100 mL flask containing 20 mL such medium, with the inoculum size of 1%, for 12 h. Then the yield of mutant human glucagon-like peptide-1 fusion protein reached 348.6 mg/L.