Abstract Regeneration is important for vegetative propagation of excellent variety, detoxification and the obtain of transgenic plant, but plant regeneration is time-consuming. Here, we found that duckweed regeneration could be enhanced by regenerating callus. Firstly, Genetic transcript regulation has been applied to study the molecular mechanism controlling regeneration. Auxin related genes have been significantly down-regulated in regenerating callus. Cytokinin signal pathway genes have been up-regulated in regenerating callus. Secondly, volatile organic compounds release has been analysised by gas chromatography/mass spectrum during the stage of plant regeneration, and 11 kinds of unique volatile organic compounds in the regenerating callus were increased. Among them, cyclohexane treatment enhanced duckweed regeneration by initiating root. Moreover, Auxin signal pathway genes were down-regulated in callus treated by cyclohexane. All together, these results provide novel mechanistic insights into how regenerating callus promotes duckweed regeneration. Graphical abstract