Hairy root clones of Rehmannia glutinosa were established via transformation with Agrobacterium rhizogenes ATCC15834. To optimize the culturing conditions for both root growth and catalpol production, effects of various combinations of seven basal media, pH, and carbon sources were examined under darkness. The fastest root growth was obtained in an SH medium containing $4\%$ sucrose (pH 5.8); the highest catalpol content ($0.54\%$ of dry weight) was achieved in a WPM medium supplemented with $4\%$ sucrose (pH 5.8). Effects of plant growth regulators and chitosan were also investigated. IAA at $2mg L^{-1}$ significantly increased root lengths and the frequency of lateral roots. Chitosan $(50 mg L^{-1});and;GA_3 (0.5 mg L^{-1})$ induced catalpol production, with contents calculated at $0.7\%$ dry weight and $0.65\%$dry weight, respectively.