Abstract:

To obtain high yield and high viability protoplasts, an efficiency protocol was presented here using leaves of in vitro shoot culture of Populus pseudo-simonii Kitag.. The yield of protoplasts depends on factors such as enzyme concentration, osmoticum concentration, incubation time and the age of leaves. The results showed that the optimal protocol for isolating protoplasts was treating 35 days aged leaves with CPW salts + 3.0% Cellulase R-10 + 0.5% Macerozyme R-10 + 0.1% Pectolase Y-23 + 0.6 mol/L mannitol + 0.6 g/L MES+ 1 g/L BSA for 8 h. Protoplast yields were up to 2.44×107 protoplasts per gram and the viability was 78.7%. This protocol that resulted in high yield and high viability of protoplasts could meet the demands of protoplast culture-based techniques.