Key Laboratory of Biomass Chemical Engineering of Ministry of Education, Department of Chemical and Biological Engineering, Zhejiang University, Hangzhou 310027, China.
Bioresour Technol. 2013 Sep;144:693-7. doi: 10.1016/j.biortech.2013.06.120. Epub 2013 Jul 5.
The cbh1 strong promoter was employed to over-express the cbh2 gene for enhancing cellobiohydrolase (CBH) production in Trichoderma reesei because CBH II component has higher specific activity than CBH I and is an important component in cellulase. The recombinant plasmid pCAMBIA1300-hph-PsCT containing strong expression cassette was constructed and transformed into T. reesei via optimized Agrobacterium-mediated transformation, producing 324 positive T. reesei transformants for the two steps of screening. Ten fast-growing T. reesei transformants were selected, amongst which C10 was found to have the highest filter paper activity 28.92±2.45 IU/mL, 4.3-fold higher than that of ZU-02, 6.71±0.79 IU/mL. C10 also has the highest cellobiohydrolase activity 122.44±7.42 U/mL, 5.4 times higher than that of ZU-02, 22.49±2.27 U/mL. The cellulase from C10 performed better (93.06±2.83%) than the one from ZU-02 in enzymatic hydrolysis because the exo-exo-synergism played a role.
采用 cbh1 强启动子过表达 cbh2 基因，以提高里氏木霉中的纤维二糖水解酶（CBH）产量，因为 CBH II 组分的比活性高于 CBH I，是纤维素酶的重要组成部分。构建了含有强表达盒的 pCAMBIA1300-hph-PsCT 重组质粒，并通过优化的农杆菌介导转化法转化到里氏木霉中，经过两步筛选，产生了 324 个阳性里氏木霉转化子。从 10 株生长较快的里氏木霉转化子中筛选出 C10，其滤纸酶活最高，为 28.92±2.45IU/mL，比 ZU-02 高 4.3 倍，为 6.71±0.79IU/mL。C10 的细胞二糖水解酶活也最高，为 122.44±7.42U/mL，比 ZU-02 高 5.4 倍，为 22.49±2.27U/mL。C10 的纤维素酶在酶解中表现更好（93.06±2.83%），因为发挥了外切-内切协同作用。
