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| Cloning of Endoglucanase Gene from Trichoderma viride and the Effect of Its Site-directed Mutagenesis on Enzyme Activity |
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Abstract Cellulose is the earth's most abundant resources of renewable resources, Cellulase enzymes and the production cost restricts the use of cellulose. In this study, a fungi G-1 was isolated from rotten wood(Populus bonatii) placed outdoor for years and indentified as Trichoderma viride through 18S rDNA sequence analysis. A endoglucanase gene(EG) (GenBank Access No.HM116999.1) was cloned from G-1 and a mutant sequence (EG-mut) was obtained by overlap primer extension method for site-directed mutagenesis of EG. Both EG and EG-mut were inserted into the secretion expression vector pPIC9K and imported in Pichia pastoris, and two strains were screened by Congo red staining and enzyme activity assay DNS method. Results showed that the activity of enzyme was respectively 20.915 and 24.110 U/mL, significantly higher than the other strains obtained in this experiment. The results indicated that some site-directed mutagenesis can reveal important functional domains of enzyme activity.
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Received: 07 January 2011
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