Abstract: In order to increase selection efficiency of tobacco breeding, we carried out an analysis of relationship between nicotine content and agronomic traits. In this study, 13 agronomic traits of 116 tobacco varieties were measured in 4 consecutive years. Correlation analysis, factor analysis, stepwise selection analysis and quadratic response surface analysis were used to explore the relationship between nicotine content and these traits. The results showed that variations in main agronomic traits of tobacco varieties were abundant with the coefficients of variation ranging from 5.64%-16.96% and the variation coefficient of nicotine content was 24.3%. Simple correlation analysis showed that nicotine content was significantly positively correlated with the internodal distance, and negatively correlated with stem girth and leaf number. Factor analysis showed that leaf length-width ration factors, leaf number factors, leaf length factors, plant height factors were largely responsible for nicotine content. Regression models, which successfully forecasted the relationships between agronomic traits and nicotine content were established by Stepwise selection method analysis. Statistical test was performed and the results showed that the correlation coefficient was significant. Furtherly, second response surface analysis revealed the dynamical relationship of nicotine content between a leaf length and middle leaf length. When bottom leaf length is 63.7 cm and middle leaf length is 58.7 cm, the nicotine content of a tobacco plant is stable and has a minimal value. Nicotine content will increase when tobacco plants have a smaller leaf length-width ratio, bigger internodal distance, less leaf number, and higher plant height. Therefore, when breeding for flue-cured tobacco varieties with different nicotine contents, breeders can use these agronomic traits as reference.