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Heterologous Expression of an Alkali and Thermotolerant Lipase from Talaromyces thermophilus in Trichoderma reesei

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Abstract

To heterologously express a Talaromyces thermophilus lipase gene in Trichoderma reesei, an efficient binary vector pChph-pCBH1sigpro-ttl which includes a newly designed cbh1 promoter and hygromycin-resistant marker was constructed. This plasmid was then transformed into T. reesei via improved Agrobacterium EHA 105-mediated transformation. After modification of co-culture conditions and enzymolysis treatment of conidia, 258 transformants were produced. A two-step screening method based on antibiotic resistance and capacity to utilize lactose and tributyrin was introduced to further select promising candidates, which would be additionally verified by PCR analysis, sodium dodecyl sulfate–polyacrylamide gel electrophoresis (SDS-PAGE), and lipase activity assay. Lipase production was carried out in shaking flasks, and the activity reached 241 IU/mL (7415.4 IU/mg) after 84-h fermentation. It was found that this lipase performed high alkali and thermostable tolerance with the optimal pH 9.5 and temperature 60 °C, and it could retain more than 70 % activity after being disposed in pH 11 or 70 °C for 1 h. This study herein would benefit the genetic engineering of T. reesei and the industrial application of this important fungal lipase.

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Acknowledgments

This work was supported by the National High-tech R&D Program (2007AA05Z401) and the Program for Zhejiang Leading Team of S&T Innovation (2011R50002) of China.

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Authors and Affiliations

  1. Key Laboratory of Biomass Chemical Engineering of Ministry of Education, Department of Chemical and Biological Engineering, Zhejiang University, Hangzhou, 310027, China

    Xu Zhang, Xueqi Li & Liming Xia

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  1. Xu Zhang
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  2. Xueqi Li
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  3. Liming Xia
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Correspondence to Liming Xia.

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Sequence alignment of the original and optimized lipase sequence (GIF 498 kb)

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Zhang, X., Li, X. & Xia, L. Heterologous Expression of an Alkali and Thermotolerant Lipase from Talaromyces thermophilus in Trichoderma reesei . Appl Biochem Biotechnol 176, 1722–1735 (2015). https://doi.org/10.1007/s12010-015-1673-4

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