In the present study, 18 karyotypes in 40 species belonging to eight subgroups of Drosophila melanogaster species group in China were analyzed. Of the 18 karyotypes, six (A, A′′, C′′′, C′′′′, D′′ and F′) were novel. In the eight subgroups, F, F′, G and H were basic karyotypes for the Drosophila ananassae subgroup, A and A′′ for D. elegans subgroup, C for D. eugracilis subgroup, C′ for D. ficusphila subgroup, C and C′ for D. melanogaster subgroup, C, C′, C′′, D, D′, D′′, E, E′, E′′ and I for D. montium subgroup, C′′′ and C′′′′ for D. suzukii subgroup, and C, C′′′ and C′′′′ for D. takahashii subgroup. In D. lucipennis, the karyotype for female was 2n=8, while that for male was 2n=7, in which one of the chromosome Ⅳ was absent. In addition, B chromosomes were found in D. kikkawai, D. lini, D. ogumai, D. pseudoananassae, and D. triauraria. The karyotypes of six undescribed species were reported in this study, namely D. sp. like elegans, D. sp. like nyinyii, D. sp. like trapezifrons1, D. sp. like takahashii, D. sp. like trapezifrons2 and D. sp. like auraria. In this paper, the karyotype of a newly recorded species D. giriensis was also determined to be 2n=8, which be-longs to the C type. Our karyotype analysis reveals diversity in karyotypes of the Drosophila melanogaster species group in China, which will provide cytological evidence for further studies on genetics and evolution of Drosophila.