Protoplast isolation in sugarcane (saccharum complex.)

The investigation was carried out on two sugarcane varieties, namely CoN -05071 and CoC-671 at the Sugarcane Tissue Culture Laboratory of Main Sugarcane Research Station, Navsari Agricultural University, Navsari during 2016-017. The current study deals with the studies on protoplast isolation such as protoplast purification, protoplast density, protoplast viability in Sugarcane (Saccharum complex). In this experiment, successful isolation of protoplast from mesophyll cells of inner two leaf whorls and callus suspension cultures of sugarcane cultivars CoC-671 and CoN-05071 was obtained. Cultivars were standardized at 1 per cent cellulase, 0.5 per cent macerozyme, 1.0 per cent pectolyase and 0.5 M manitol. Maximum protoplast yield was observed at 600 rpm in callus suspension cells (68%) at 2 hrs of incubation period. Maximum density of protoplast was observed in callus suspension derived protoplast culture ((2.0×104, 2.2×104) in both the varieties at 2 hrs of incubation periods. Highest protoplast viability observed at 2.2x104 density level of callus suspension origin protoplast. In this study 2 hrs of incubation is optimal time for protoplast isolation from mesophyll cells and callus suspension cells of sugarcane cultivars.