Abstract: 【Objective】The objective of this study is to establish an efficient system of protoplast regeneration for further developing protoplast fusion and transformation in cassava. 【Method】 Protoplasts were isolated from suspension cultures derived from friable embryogenic callus (FEC) of cassava genotype TMS60444. The highest protoplast yield obtained was 3.5×106 protoplasts/g fresh weight. Viabilities of the protoplasts assessed by the fluorescein diacetate (FDA) were approximately 90%. Protoplasts were cultured in TM2G medium with liquid thin layer culture at densities of 5×105 p/mL or 2×105 p/mL. During the first 30 d, the medium was refreshed by 0.3 mol•L-1 TM2G fresh medium every 10 d. After that, the medium was refreshed by 0.25 mol•L-1 TM2G fresh medium every 10 d. After cultured for 45 d, calli of 1-2 mm were picked out and separately developed into embryos on MSN medium, into mature embryos on CMM medium, into shoots on CEM medium and into roots on MS medium. 【Result】It was showed that all protoplasts cultured at density of 5×105 p/mL developed into compact calli (could develop into embryos), protoplasts cultured at density of 2×105 p/mL developed into compact calli and vacuolar calli (could not develop into embryos). A total of 1 479 compact calli were picked out and developed into 757 cotyledon embryos and regenerated 186 plants in the experiment. 【Conclusion】The yield and viability of isolated protoplasts had been greatly increased, the bottleneck of predecessors mentioned was improved, and the efficiency of plant regeneration from protoplasts was promoted. 

Key words: cassava, friable embryogenic callus (FEC), protoplast, plant regeneration