Chinese soft-shelled turtle (Pelodiscus sinensis) is an important freshwater breeding species in East and Southeast Asia. The most external differences among different strains are skin pigmentation and blotches, but the mechanism of their formation is unknown. To investigate the mechanism of skin pigmentation diversity in Chinese soft-shelled turtle, three unique phenotypes (Golden strain, Japanese strain and Qingxi black strain) were selected to observe the chromatophores, and the transcriptome and metabolome were compared. The results showed the presence of melanophores, xanthophores and iridophores in the skin of all three strains. However, there were differences in the composition and organelles of chromatophores. The upregulation of gene asip and elevated accumulation of glutathione and cysteine in Golden strain resulted in the production of pheomelanin, leading to yellow pigmentation in the skin. The upregulation of gene dct and elevated accumulation of indole-5, 6-quinone in Qingxi black strain resulted in the production of eumelanin, which made the skin appear black-brown. Japanese strain produced eumelanin and pheomelanin, however, the presence of melanosomes resulted in skin darkening and appeared dark green pigmentation. The white blotches of Gloden strain and Japanese strains were formed through increased production of iridophores, while the black blotches of Qingxi black strain were formed by inducing migration and apoptosis of melanophores within iridophores. In conclusion, the distinctive characteristics of the skin in three strains resulted from combination of melanin type, abundance of melanosomes and interaction between melanophores and iridophores. These results provided valuable reference for investigating the mechanisms underlying skin pigmentation diversity in Chinese soft-shelled turtle and other vertebrates.