Isoflavone contents were analyzed by HPLC methods in 100 representative soybean cultivars from different regions in this study. The results showed that all of 100 cultivars contained six of major isoflavone components in soybean seeds, including daidzin (D), glycitin (GL), genistin (G), malonyldaidzin (MD), malonylglycitin (MGL) and malonylgenistin (MG), and there were significant difference on the isoflavone content among ecological regions and varieties, which explained the abundant genetic variation of isoflavone content in the soybean germplasm. The average of total isoflavone content (2465.48 ug/g) in soybean cultivars from south region was the highest, and that from north region was the lowest (1705.89 ug/g). Coefficient of variation (CV) for the six major isoflavone components was ranged from 33.44% to 52.03%. Correlation analysis showed that there was significantly positive correlation among the isoflavone components, in which correlation coefficient between total isoflavone content and malonylgenistin was the maximum (r = 0.958 **); A significantly negative correlation was presented between total isoflavone content and oil content (r = -0.323 **). Two soybean cultivars with high isoflavone content (cv. Pingdingheidou, 4459.91ug/g and cv. PI-567479, 4073.95 ug/g), and two cultivars with low isoflavone content (cv. Chasedou, 857.74 ug/g and cv. Mufeng1, 922.82 ug/g) were selected for soybean breeding and genetic study on the isoflavone.