Brassinosteroids (BRs) are natural plant growth regulators required for plant growth and development. Physiological studies have demonstrated that exogenous BRs induced the elongation of coleoptiles and mesocotyls in maize. However, little is known about the function of endogenous BRs in maize or other grass species. The Arabidopsis DWF1/DIM is a bifunctional protein which catalyses both the △24 isomeration, and subsequently, the △24(25) reduction in BRs biosynthesis pathway. To better understand the role of DWF1 of monocot plants in BRs biosynthesis, we cloned a maize gene, ZmDWF1, which has extensive sequence similarity to Arabidopsis DWF1. ZmDWF1 is predicted to encode a 562-amino-acid protein that has 95 and 86% similarity with rice OsDWF1 and Arabidopsis DWF1, respectively. Southern blot result showed that one copy of ZmDWF1 is present in maize genome. Northern blot analyses revealed that expression of ZmDWF1 is temporally and spatially regulated during development. Transgenic maize plants with double-stranded ZmDWF1 RNA were generated. Some of them have dwarfed phenotype varying in severity. Western blot results also demonstrated the expected cosuppression of ZmDWF1 gene in transgenic plants.