Sorghum (Sorghum bicolor) roots exude a potent bioherbicide sorgoleone. Previous work indicates that sorgoleone is produced in living root hairs. We have developed a mist system that resulted in abundant production of root hairs exuding sorgoleone and a mat system that significantly inhibited root hair development and consequently sorgoleone production. Applying Ag+ (an ethylene action inhibitor) at 1.2 mM to the seedlings grown in the mist system also inhibited root hair formation and elongation. Hypoxic conditions in the mist system did not result in the inhibition of root hair growth as compared to the standard air atmosphere (20.8% O2). Applying ethephon (an ethylene-releasing agent) at 0.031 mM to the roots of seedlings grown in the mat system with water running at 1 ml/min reversed the inhibition of root hair development by water movement. These results indicate that either water movement or ethylene can be utilized to manipulate root hair development and sorgoleone production in sorghum seedlings. It is hypothesized that water movement reduced the local ethylene concentration on the root surface and consequently inhibited root hair development of sorghum seedlings grown in the mat system.