Breeding and application of disease resistant varierties is the preferred method to control Verticillium wilt diseases, it is important to broaden the genetic base of upland cotton for continuous genetic improvement of disease resistance. The objectives of this study were to evaluate 107 upland cotton main breeding germplasm includes introgressed lines from G. barbadense were identified in Verticillium dahliae nursery for three years repeat identification experiments. The results showed that only 8 disease-resistant germplasm were screened, accounted for only 7.5% of 107 accessions; disease-tolerant cultivars or lines were 20 accessions, were 18.7%; The identified results indicated that it is scarcity of sources of disease resistant germplasm can be used as breeding parent at present. So it is necessity to research on disease-resistant genetic mechanism and collaborative improved relations with other economic characters, and provided theoretical guidance for cotton resistance breeding. The analyses suggest that distant hybridization breeding was a powerful way of creation of germplasm resistance to verticillium wilt in upland cotton.