High frequency one-step gene replacement in Trichoderma reesei. II. Effects of deletions of individual cellulase genes |
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Authors: | Pirkko L Suominen Arja L Mäntylä Taina Karhunen Satu Hakola and Helena Nevalainen |
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Institution: | (1) Research Laboratories, Alko Ltd, P.O.B. 350, SF-00101 Helsinki, Finland;(2) Fazer Chocolates Ltd, P.O.B. 4, SF-00941 Helsinki, Finland;(3) Present address: School of Biological Sciences, Macquarie University, 2109 Sydney, N.S.W., Australia |
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Abstract: | Four cellulase genes of Trichoderma reesei, cbh1, cbh2, egl1 and egl2, have been replaced by the amdS marker gene. When linear DNA fragments and flanking regions of the corresponding cellulase locus of more than 1 kb were used, the replacement frequencies were high, ranging from 32 to 52%. Deletion of the major cellobiohydrolase 1 gene led to a 2-fold increase in the production of cellobiohydrolase II; however, replacement of the cbh2 gene did not affect the final cellulase levels and deletion of egl1 or egl2, slightly increased production of both cellobiohydrolases. Based on our results, endoglucanase II accounts for most of the endoglucanase activity produced by the hypercellulolytic host strain. Furthermore, loss of the egl2, gene causes a significant drop in the filter paper-hydrolysing activity, indicating that endoglucanase II has an important role in the total hydrolysis of cellulose. |
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Keywords: | Trichoderma reesei Gene replacement Homologous recombination Cellulases Regulation |
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