Flower-like CuO nanostructures with porous nanosheets were synthesized on alumina tube using a hydrothermal method without using any surfactant. The flower-like CuO was comprised of interconnected nanosheets with a uniform thickness of 60 nm. These nanosheets had numerous nanoscale pores with diameters ranging from 20 nm to 160 nm. The copper complex ions of [Cu(NH3)4]2+ and NH3 were identified to be critical for the formation of these flower-like CuO nanostructures with porous nanosheets during the hydrothermal process. Gas sensor to hydrogen sulfide (H2S) based on these porous flower-like CuO nanostructures exhibited high sensitivity, good reproducibility and long-term sensing stability when tested at room temperature of 25 °C. The gas sensor also showed a remarkably high sensing selectivity to the H2S gas. The gas sensing mechanism was investigated, and the formation of CuS on the surface of CuO nanostructure was identified to be critical for H2S sensing.