Abstract: The female loach (Misgurnus anguillicaudatus) generally have a larger size compared to males. The application of artificially induced gynogenesis in the cultivation of strains presents great potential. In this study, the starting time of heat shock of artificially induced gynogenesis was determined by cytological observation after cell fluorescence staining. The artificially induced gynogeny embryos were further identified by using karyotype and flow cytometric analysis. The results showed that artificial gynogenesis of loach was successfully induced by inactivated common carp sperm. Cytological observation of fertilization showed that the second polar body was observed at 3—5min after insemination. The optimal starting time of heat shock treatment at 41℃ was found to be 3.5min after insemination, followed by a 2min heat shock, resulting in an optimal hatchability rate of 10.26%. By analyzing the development of wild-type loach embryos, artificially induced gynogenetic loach embryos, loach × carp hybrid embryos and haploid loach embryos, it was found that some gynogenetic embryos developed normally, while the hybrid and haploid embryos showed obvious developmental disorders. Karyotype and flow cytometric analysis of the gynogenetic loach embryos further demonstrated that treatment with the heat shock parameters obtained in this study resulted in 64.71% diploid gynogenetic offspring. In this study, the optimized induction parameters of loach meiogynogenesis were obtained through the cytological analysis, which could provide a technical basis for the artificially induced meiogynogenesis in the cultivation of all-female loach strains.