Biotechnology and Bioprocess Engineering 2019; 24(3): 500-506  
Elevating Limonene Production in Oleaginous Yeast Yarrowia lipolytica via Genetic Engineering of Limonene Biosynthesis Pathway and Optimization of Medium Composition
Bo-Qian Cheng1,†, Liu-Jing Wei1,†,*, Yu-Bei Lv1, Jun Chen1, and Qiang Hua1,2
1State Key Laboratory of Bioreactor Engineering, East China University of Science and Technology, Shanghai 200237, China
2Shanghai Collaborative Innovation Center for Biomanufacturing Technology (SCICBT), Shanghai 200237, China
Correspondence to: Liu-Jing Wei
Tel: +86-021-64252147; Fax: +86-021-64250972

These two authors contributed equally to this work.
Received: December 27, 2018; Revised: March 13, 2019; Accepted: March 24, 2019; Published online: June 30, 2019.
© The Korean Society for Biotechnology and Bioengineering. All rights reserved.

This is an Open Access article distributed under the terms of the Creative Commons Attribution Non-Commercial License ( which permits unrestricted non-commercial use, distribution, and reproduction in any medium, provided the original work is properly cited.
As an alternative terpenoid producer, non-conventional oleaginous yeast Yarrowia lipolytica was chosen for limonene production. Y. lipolytica can give high biomass yield and metabolize a broad range of substrates such as glycerol, alkanes, fatty acid, fats, and oils. As previously reported, optimization of limonene synthesis pathway and mevalonate (MVA) pathway leads to the accumulation of 112-fold higher limonene as compared to an initial strain. In this study, we introduced an additional copy of limonene synthesis gene (LS), which resulted in an increase of limonene production. This engineered strain was used to carry out further optimization study. Amongst all the carbon sources tested, the highest level of limonene production was obtained from glycerol, and citrate was selected as an auxiliary carbon source. In fed-batch fermentation with an optimized medium, the engineered strain was found to produce 165.3 mg/L limonene, which corresponds to the highest yield till date for the production of limonene in Y. lipolytica.
Keywords: limonene, Yarrowia lipolytica, carbon source, fed-batch fermentation

This Article