Recombinant human growth hormone (r-hGH) can be produced by recombinant Escherichia coli a main host for recombinant protein productions. In order to improve the productivity of r-hGH, an orthogonal (L16(43×22)) experiment was applied to optimize the best culture conditions for r-hGH production by flask cultures of E. coli BL (21) harboring a new constructed plasmid (pEHUb-hGH). Based on the results of primary tests, five factors such as culture medium, culture volume, induction starting time, r-hGH expression time, and IPTG concentration were chosen for this present investigation as themain factors to influence the r-hGH expression. The optimum culture conditions were determined as follows: culture mediumas themodified TB, culture volume 50mL, induction starting time 3 h, r-hGHexpression time 7 h, and IPTG0.3mmol/L.Under this optimized conditions, the r-hGH productivity could reached 18.72 mg/L·h in flask culture. A high r-hGH concentration of 4.3 g/L, resulting in a high productivity of 215 mg/ L·h, could be obtained in the fed-batch culture of recombinant E. coli.