Journal
BIORESOURCE TECHNOLOGY
Volume 110, Issue -, Pages 572-577Publisher
ELSEVIER SCI LTD
DOI: 10.1016/j.biortech.2012.01.129
Keywords
Cellulase; T-DNA insertional mutagenesis; Trichoderma reesei; Screening
Funding
- National Basic Research Program of China [2011CB707403]
- National Natural Science Foundation of China [30800024, 30970026]
- Excellent Young and Middle-Aged Scientist Award Grant of Shandong Province of China [BS2011SW017]
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A T-DNA-tagged mutant library created by Agrobacterium-mediated transformation (AMT) was assayed for improvement of cellulase production. After 96-well plate screening for rapid growth on cellulose substrates followed by plate-clearing zone assay, three putative mutants, TA-32, TB-87 and TE-6, with enhanced cellulolytic ability were isolated, exhibiting 38%, 51% and 31% increase in total cellulase activity than the parental strain QM9414, respectively. Endoglucanase, cellobiohydrolase and beta-glucosidase activities as well as the hydrolysis efficiencies of the mutants were also improved. Moreover, T-DNA was shown to be integrated at a single site in the genomes of TA-32 and TE-6 while inserted at two copies into the genome of TB-87. Further, the sequences flanking the T-DNA insertion sites were successfully rescued, demonstrating the increased utility of T-DNA insertional mutagenesis for improvement of cellulase production as well as subsequent identification of the tagged genes relevant to cellulolytic ability. (C) 2012 Elsevier Ltd. All rights reserved.
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