Abstract A new protocol has been developed for the highly efficient somatic embryogenesis and plant regeneration of 10 recalcitrant Chinese cotton cultivars. Calluses and embryogenic calluses were induced on MSB1 medium containing the optimal combination of indolebutyric acid (IBA; 2.46 μM) and kinetin (KT; 2.32 μM). Up to 86.7% of embryogenic calluses differentiated into globular somatic embryos 2 mo. after culture on MSB2 medium containing double KNO3 and free of growth regulators. Up to 38.3% of the somatic embryos were converted into complete plants in 8 wk on MSB3 medium with l-asparagine (Asn)/l-glutamine (Gln) (7.6/13.6 mM). The plants were successfully transferred to soil and grew to maturity. With the protocol described here, we have obtained hundreds of regenerating plantlets from 10 recalcitrant cultivars, which is important for the application of tissue culture to cotton breeding and biotechnology.