Abstract: In order to achieve the plantlet sugar-free culture in open space, a sugar-free culture system with environmental control device was set up. The air in system culture container flowed through the pipeline to air-processing equipment. The environment cleanliness in culture container was ensured and the spreading bacteria was cut off. The system cultivated the plantlet worked with carbon dioxide, supported by perlite and provided by artificial light and nutrition solution on time. Temperature and relative humidity was controlled using upper and lower limits, CO2 concentration was controlled using fuzzy-integral method in culture container. The test showed that the temperature was controlled at 20～30℃ and relative humidity at 72%～76% during illumination, the temperature at 14℃ or more and relative humidity at 80%～85% during dark. The control of CO2 concentration had no overshoot, the rise time was 30min, the steady-state error was 43μmol/mol, and therefore in this way the culture quality of chrysanthemum plantlet was better than that in a test tube.