Circular RNA (circRNA) is a regulatory class of long, noncoding RNA found in both plant and animal kingdoms. The profile and characterization of circRNA in cotton species remains to be explored. Here, using 24 rRNA-depleted RNA-seq libraries of putative diploid progenitors of Gossypium spp., Gossypium arboreum and Gossypium raimondii, their interspecies hybrid (F ) and allotetraploid Gossypium hirsutum, 1041, 1478, 1311, and 499 circRNAs were identified in each cotton species, respectively. A prevalence of 23 exon-circRNAs contain noncanonical GT/AG signals, and only ~ 10% of exon-circRNA is associated with reverse complementary intronic sequences. This result implies that plants employ a method of circRNA splicing distinct from that of animals. In addition, 432 circRNAs are stably expressed in multiple cotton species.