Production and regeneration conditions of Trichoderma koningii protoplast

To provide a suitable fungal material for protoplast-fusion and mutation-breedin g of Trichoderma koningii,conditions of the fungal protoplast preparation and regeneration including mycelium growth time,the content of cellulase,temperature and time for reaction and the composition and concentration of various osmotic stabilizers were investigated.The optimal conditions for protoplast preparation of T koningii were as follows: mycelium growth time 22 h,the content of cellulase 2.0%,reaction time 2.5 h and reaction temperature 30℃.The optimal osmotic stabilizer in the snail enzyme solution and the osmotic pressure stabilizer in the regeneration culture medium were 0.6 mol/ L NaCl and 0.8 mol/L sucrose,respectively.Microscopical observation indicated that the protoplasts of T.koningii were largely released from the apex and mycelium position.One modes of forming irregular outshoot were found to regulate regeneration of T.koningii.An optimal concentration of T.koningii protoplasts can be achieved using the preparation system.