Author:
Li Ya-Wen,Yang Cai-Ling,Shen Qi,Peng Qian-Qian,Guo Qi,Nie Zhi-Kui,Sun Xiao-Man,Shi Tian-Qiong,Ji Xiao-Jun,Huang He
Abstract
Non-homologous end-joining (NHEJ)-mediated random integration in Yarrowia lipolytica has been demonstrated to be an effective strategy for screening hyperproducer strains. However, there was no multigene assembly method applied for NHEJ integration, which made it challenging to construct and integrate metabolic pathways. In this study, a Golden Gate modular cloning system (YALIcloneNHEJ) was established to develop a robust DNA assembly platform in Y. lipolytica. By optimizing key factors, including the amounts of ligase and the reaction cycles, the assembly efficiency of 4, 7, and 10 fragments reached up to 90, 75, and 50%, respectively. This YALIcloneNHEJ system was subsequently applied for the overproduction of the sesquiterpene (-)-α-bisabolol by constructing a biosynthesis route and enhancing the flux in the mevalonate pathway. The resulting strain produced 4.4 g/L (-)-α-bisabolol, the highest titer reported in yeast to date. Our study expands the toolbox of metabolic engineering and is expected to enable a highly efficient production of various terpenoids.
Funder
National Key Research and Development Program of China
Natural Science Foundation of Jiangsu Province
Subject
Biomedical Engineering,Histology,Bioengineering,Biotechnology
Cited by
16 articles.
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