The growth information and transpiration information of plantlets in vitro were acquired by image processing and utilization of the law of mass conservation, respectively. The Logistic, linear and nonlinear hyperbolic equations were used for fitting the relationship of biomass of plantlet in vitro and culture time, water loss and culture time, transpiration water loss and biomass of plantlet in vitro. The models of growth prediction, transpiration of water loss and water using efficiency of plantlet were established. The transpiration rate and water using efficiency of plantlet in vitro were calculated. The transpiration rate and water using efficiency of Atractlodes lancea plantlets in vitro at different nitrogen levels in the medium and subculture generation were measured. The transpiration was strong with less subculture generation and higher nitrogen content in culture medium. 