Abstract: In order to establish an efficient and large-scale cotton transgenic technology system to solve the problems, such as the conversion of cotton genotypes restricted, low conversion efficiency and the conversion rate of seedlings deformities etc., by using the agrobacterium-mediated gene transformation method, the Bt gene was introduced into embryogenic callus of xinhai16 (Xinjiang Sea Island cotton). The study aims to improve Sea Island cotton genetic transformation efficiency and establish a relatively efficient regeneration of transgenic plants seedling technology system. The results showed that: the callus subculture on callus proliferation resistant medium containing cefotaxime, following three times, each subculture medium containing cephalosporins had a concentration gradient, or we could let callus subculture 3 times, each time subculture containing 500 mg/L cefotaxime resistant callus proliferation medium MS₂. Compared with the latter, the speed of resistant callus proliferation of the former is faster. What’s more, adding 50 mg/L Kan per 60 days, the seedling rate of transformed seedlings reached the maximum. Reducing the level of NH₄ /NO₃^- had an effective role in improving the rate of somatic embryogenesis. Selecting subculture callus after 90 days, embryo induction rate could reach the maximum value. After subculture time of 90 days, embryoid induction rate reached the maximum; more than 180 days of subculture time, the callus could be abandoned. After transforming, the cotyledon abnormal embryos were obtained with a strong ability to regenerate, its secondary embryos could produce normal transgenic plants again by cultivating and this approach could effectively shorten the conversion period.