Abstract: In order to broaden the genetic variation and improve the resistance toVerticillium wilt of cotton varieties in Yangtze River Basin, a comprehensive population was built by 19 cotton varieties from Yellow River Basin through mixed hybridization, 37 elite lines were derived through breeding selection at Yangtze River Basin and 14 traits were investigated by two years’field experiment. The results showed that the derived lines had the greatest variation in the resistance toVerticillium wilt , and then was in yield traits, the lowest variation was found in fiber quality traits. Clustering analysis was performed by SSR markers, the 37 derived lines showed large genetic variation and could be dispersed in different cluster groups, which showed a clear distinction from the cotton varieties from the Yangtze River Basin. The factor analysis was performed on the investigated traits of the derived lines, the first 7 common factors contributed to the total variance up to 83.08%, fiber quality factor, plant type factor, yield factor and disease index factor had the contribution rate of 28.45%, 15.50%, 12.07% and 8.87%, respectively.