High efficient expression of cellobiase gene from Aspergillus niger in the cells of Trichoderma reesei.
The cellobiase gene from Aspergillus niger was cloned and connected with the strong promoter Pcbh1 from Trichoderma reesei to construct a recombinant plasmid pHB9 with the hygromycin B resistance marker. The plasmid was transformed into conidia of T. reesei using the modified PEG-CaCl(2) method. Main factors effecting the transformation were discussed and about 99-113 transformants/μg DNA could be obtained under optimal conditions. It was found that the molecular mass of the recombinant cellobiase was about 120 kDa by SDS-PAGE analysis. The activity of cellobiase could reach 5.3 IU/ml after 48 h fermentation, which was as high as 106 times compared with that of the host strain. Meanwhile, the filter paper activity of recombinant T. reesei was 1.44-fold of the host strain. Saccharification of corncob residue with the crude enzyme showed that the hydrolysis yield (84.2%) of recombinant T. reesei was 21% higher than that (69.5%) of the host strain.
Department of Chemical Engineering and Bioengineering, Zhejiang University, Hangzhou 310027, China.
Bingbing
B
Wang
Liming
L
Xia
D001234:Aspergillus niger / Q000187:drug effects / Q000201:enzymology* / Q000235:genetics*; D002480:Cellulase / Q000096:biosynthesis; D015825:Chromosomes, Fungal / Q000235:genetics; D004271:DNA, Fungal / Q000235:genetics; D004591:Electrophoresis, Polyacrylamide Gel; D015870:Gene Expression / Q000187:drug effects; D005800:Genes, Fungal / Q000235:genetics*; D006868:Hydrolysis / Q000187:drug effects; D010957:Plasmids / Q000235:genetics; D011092:Polyethylene Glycols / Q000494:pharmacology; D016133:Polymerase Chain Reaction; D011523:Protoplasts / Q000187:drug effects / Q000378:metabolism; D011995:Recombination, Genetic / Q000235:genetics; D015203:Reproducibility of Results; D013997:Time Factors; D014170:Transformation, Genetic / Q000187:drug effects; D014242:Trichoderma / Q000166:cytology* / Q000378:metabolism*; D003313:Zea mays / Q000378:metabolism; D001617:beta-Glucosidase / Q000235:genetics* / Q000378:metabolism
D004271:DNA, Fungal; D011092:Polyethylene Glycols; D001617:beta-Glucosidase; D002480:Cellulase