영지 원형질체 분리를 위한 대체 세포벽 분해 효소 탐색과 최적 조건 설정

RESEARCH ARTICLE

영지 원형질체 분리를 위한 대체 세포벽 분해 효소 탐색과 최적 조건 설정
Exploration of alternative cell wall lysing enzymes and optimization of conditions for Ganoderma lucidum protoplast isolation

영어: Recently, active research in Korea and worldwide has begun to focus on gene function and cultivar development using gene editing tools. This research, in addition to studies on edible mushroom, aims to enhance the physical and biochemical characteristics of mushrooms for applications in materials and substance production. For these studies, efficient isolation of protoplasts from the target mushroom is critical. However, several commercial cell wall-lysing enzyme cocktails, including Novozyme234, Glucanex, and Lysing enzymes, have recently been discontinued. In this study, we aimed to identify alternative enzyme systems to replace the discontinued cell wall-lysing enzymes for stable isolation of protoplasts from Ganoderma lucidum. To select an optimal osmotic buffer, enzyme function in 0.6 and 1.2 M Sorbitol, Sucrose, Mannitol, and KCl was assessed. The effect of reaction time was also evaluated. Protoplast isolation efficiency of each alternative enzyme was tested using lysing enzymes from Trichoderma harzianum, Chimax-N, and Yatalase, either individually or in combination. This matrix of studies identified enzymes and optimal conditions that could replace the discontinued lysing enzymes.

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