Abstract: The study aims to clarify the effects of float seedling tray size, microbial agent and transplanting leaf age on growth, yields and quality of flue-cured tobacco in field. A three-factor experiment was conducted with two size of seedling trays (136 holes or 200 holes), with or without microbial agent, and two transplanting leaf age of 8 leaves or 10 leaves of Yunyan87. Growth indexes of root and aboveground, dry matter accumulation and root-shoot ratio, chemical compositions and economic characters of flue-cured tobacco were determined. Effects of seedling tray size, microbial agents, transplanting leaf age and their interactions on growth, yields and quality of flue-cured tobacco were estimated with the value of η². Results showed that increasing the size of float seedling tray, adding microbial agents and delaying transplanting could promote the growth of root and above-ground, increase dry matter accumulation, and improve the yields and quality of flue-cured tobacco. The contribution rates of seedling tray size, microbial agent, transplanting leaf age and their interactions to the growth of tobacco root, achieved 28.92%, 23.17%, 23.64% and 24.27%, respectively. While those to aboveground growth were 27.07%, 27.64%, 31.00% and 14.29%, respectively; to dry matter accumulation, were 25.66%, 28.61%, 37.94% and 7.79%, respectively; to chemical compositions, were 25.66%, 28.61%, 37.94% and 7.79%, respectively; to economic characters were 25.66%, 28.61%, 37.94% and 7.79%, respectively. In conclusion, increasing the size of float seedling tray with microbial agents to cultivate long leaf-aged seedlings of 10 leaves, can promote early growth and rapid development of flue-cured tobacco, enhance yields and quality of tobacco leaves. The result has a certain extension value in tobacco-rice growing areas in South China.