The conditions of preparing Aspergillus niger protoplasts by enzymolysis

Mutation efficiency of microbe can be improved by mutation of protoplasts.The preparation conditions of Aspergillus niger protoplast including mycelium incubation time,osmotic stabilizer,enzymes composition,enzymatic reaction time and metal ions were investigated,and the formation procedure of A.niger protoplast was observed under the phase contrast microscope.The optimal conditions were determined that the mycelia incubated for 4dand treated with 0.5% snailase and0.5% cellulase for 3hin 0.6 mol/L NaCl osmotic stabilizer.The result showed that most suitable regeneration media contained yeast 0.2%,peptone 0.5%,NaCl 0.2%,starch 1%,CaCl20.1% and MgSO4·5H2O 0.1%.Calcium ions could improve the regeneration of A.niger protoplast more effectively than the others.