Protoplast preparation and regeneration conditions of Penicillium Q5 and Bacillus subtilis k3

The optimum conditions for the formation, regeneration and deactivation of protoplasts of Penicillium Q5 and Bacillus subtilis K3 were studied. The high-quality protoplasts of Penicillium Q5 and Bacillus subtilis K3 have been prepared by using enzymatic hydrolysis method. Through the tests, the conditions including microbial growth, the content of cellulase and time for reaction and the optimal inactivated time were optimized. The amount of protoplast from Penicillium Q5 gained a maximum, 7.35×106cfu/mL, and the corresponding above used values were 0.5% Glycolic, 10% sugar, 10 mg/mL lysozyme and snail enzyme(1 ︰ 1), 3 hours, and 35 ℃. While the product of protoplast preparation and regeneration from Bacillus subtilis K3 reached the top, and the amount of protoplast from K3 maximized 1.46×107cfu/mL, and the corresponding above used values were 4 U/mL penicillin, 1 mg/mL lysozyme, 1 hours, and 35 ℃. The period of the optimal heat inactivation in 65 ℃ was 1 hour for Penicillium Q5, while 2.5 hours for Bacillus subtilis K3. And the time of ultraviolet inactivation was 3 min for Penicillium Q5, and 5 min for Bacillus subtilis K3. The findings provided a suitable material for protoplast fusion between Penicillium Q5 and Bacillus subtilis K3, which products high-yield cellulase with resistance.