Polyethylene glycol is the most commonly used chemical in protoplast fusion as it induce reproducible high frequency fusion accompanied with low toxicity to most cell types. A mixture of pactinase or macrozyme and cellulase is used for the isolation of protoplast as these enzymes degrade the cell wall or plant cell. During cryopreservation the gemiplasm frozen and stored in liquid nitrogen at 196∘C for long time. DMSO is used as cryoprotectant in cryopreservation. Cryoprotectants prevent the formation of large ice crystals in cells and protect them from toxic solution effect due to water loss from the cytoplasm.