Abstract: With the breeding goal of special flavor, a new flue-cured tobacco cultivar with flower flavor, Zhongyantexiang 301 was selected from EMS-mutagenized Zhongyan 100, and bred through pedigree method in which descendants were continuously self-bred. The new variety was certified by the National Tobacco Variety Examination and Approval Committee in September, 2020. The variety comparison test showed that Zhongyantexiang 301 grew moderately at the early stage and gradually became stronger at the late stage, with narrow and long waist leaves and thick main stem. The main economic characters were better than Zhongyan 100. As for resistance to diseases, Zhongyantexiang 301 was resistant to brown spot disease, and medium resistant to tobacco mosaic virus (TMV) and black shank, and suitable for planting in medium-high fertility tobacco fields, like the Huanghuai area, southwest China and central China. The results of tobacco quality evaluation showed that the oil of appearance quality, reducing sugar, total sugar and ratio of total sugar to total alkaloid were higher than those of Zhongyan 100, the average score of sensory quality evaluation was higher than Zhongyan 100. In summary, Zhongyantexiang 301 had a special flower flavor, also improved the main economic characters and disease resistance.