Paclobutrazol (PAC), as a commonly used plant regulator, has the important function of improving the plant’s stress resistance. Sequoia sempervirens Endl. is a large caliber, fast-growing timber species and one of the world’s five major landscaping tree species. This study researched the effects of spraying PAC on the growth and photosynthetic activity of S. sempervirens seedlings. The results showed the plant height and the plant crown diameter were decreased significantly with the increase in PAC concentration (500 – 3,000 mg·dm−3). However, the ground shoot diameter, net photosynthetic rate, stomatal conductance, inter-cellular CO2 concentration, transpiration rate, actual photochemical quantum yield, and photosynthetic electron transport rate all showed the trend of increasing first reaching the maximum at 2,000 mg·dm−3 and then decreased. Water use efficiency showed the opposite trend. Spraying 2 times was better than once. The conclusion suggests that PAC can protect the photosynthetic activity and improve the resistance of S. sempervirens seedlings under natural cooling and draught in autumn and winter.