Abstract: To accelerate the commercial use of new flue-cured tobacco variety Yunyan97, field experiment was performed to study the effect of different nitrogen fertilizer rate on the nitrogen uptake and utilization, and the growth, yields and quality of Yunyan97. The results showed that the nitrogen content, absorption, distribution and transport were affected by nitrogen rate. With the increase of nitrogen rate, the proportion of nitrogen distribution in lower leaves and cutters declined, and the contents of nicotine and nitrogen of cured leaves from the counterpart increased, while the content of total sugar, reducing sugar and the ratio of sugar to nicotine decreased. The leaf position, the nitrogen application rate and their interaction affect the contents of chemical components. It is suggested that the optimal nitrogen application rate of Yunyan97 is 8 g N per plant, considering the factors of yields, production value and chemical components of tobacco leaves.