• 作者： 王記慧; 張立雪; 許宗雄
• 作者服務機構： 國立清華大學生命科學研究所
• 中文摘要： Trichoderma 屬黴菌為高活性纖維素水解?的重要生產菌。由於其具備不明確的性階段，進一步菌種改良乃有賴應
  用如原生質胞融合或重組DNA等遺傳操作技術，故高效率原生質胞製備方法的建立極為重要。本研究利用酵素 Driselase
  或由菌種自身培養液製備之酵素對 Trichoderma koningii G-39 之發芽管進行處理而可於約一個小時獲取高產量之原生質
  胞。探討原生質胞形成之效率之因素發現，於酵素水解作用方面，發芽管比菌絲有較高的感受性，可能因發芽管之細胞
  壁含量較少之故；而於原生質胞之產量與其穩定性方面，山梨糖（0.8M）顯為較合適之滲透壓支持劑，另原生質胞之產量
  亦直接與發芽管與酵素濃度成正比。進一步酵素活性測定顯示此二細胞壁水解系統之主要酵素活性為β-1,4-聚葡糖?，
  β-1,3-聚葡糖?，與芳基-β-醣??。另 G-39 酵素亦含有幾丁質?。至於原生質胞的再生過程則經觀察乃依如下順序進
  行：(1)先形成不規則狀之細胞，（2)整個細胞增大，(3)真菌絲形成。由本法製備之原生質胞百分之九十以上可於再生培養
  基中再生並形成菌落。t
• 英文摘要： Protoplasts from Trichoderma koningii G-39 were prepared by enzymatic hydrolysis of germ tube suspensionsutilizing Driselase or an enzyme preparation from its own culture filtrate. The formation of protoplast was almostcomplete after 1.5 hours of enzyme treatment. The efficiency of protoplast formation was dependent on a numberof factors. The germ tube was far more susceptible to enzymatic lysis than the hypha was, apparently due to thesmaller amount of cell wall in the germ tube. Sorbitol (0.8 M) appeared to be the more effective osmotic support inregard to protoplast yield as well as stability in the lytic enzyme solution. The yield of protoplasts was directlyproportional to the germ tube and enzyme concentrations. Enzymatic activity measurements indicated that the mostprominent enzymatic activities of both lytic systems were β-(1,4)-glucanase, β-(1,3)-glucanase and aryl-β-glucosidase.In addition, chitinase was present in the lytic enzyme preparation from T. koningii G-39. Regeneration of the proto-plasts was observed to proceed in the following sequence: 1) production of irregular-shaped cells followed by 2) cellenlargement and, finally, 3) true hypha formation. More than 90% of the protoplasts prepared by the describedprocedures can be regenerated and form colonies in the regeneration medium.
• 中文關鍵字： Cellulase; Trichoderma Koningii; Driselase; Protoplast; Cell Wall
• 英文關鍵字： --