Yuhui Cui, Duo Liu, Huimin Xue, Mingshan Li, Wenhong Guo, Cuiqin Huang, Xintian Zheng, Jichao Yang, Hong Liu, Huifang Yin, Hanjie Wang
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Metabolic Engineering of Yarrowia lipolytica with Massive Gene Assembly and Genomic Integration.
Synthetic biology has developed varied strategies of one-pot multigene assembly and genomic integration to promote the genetic engineering of the chassis. However, such strategies for engineering oleaginous yeast Yarrowia lipolytica is lacking, given the current stage that at most 5 exogenous genes (around 13 kb) can be assembled and integrated into the genome at once. Here, we developed a strategy of massive gene assembly and integration in the Y. lipolytica genome. As a proof of concept, dozen-gene assembly (more than 30 kb) and integration were achieved stably and reproducibly, and a Y. lipolytica chassis containing a total of 35 exogenous genes (a sum of 93.5 kb) was constructed. The introduction of massive genes modulated the synthesis of lycopene, a heterologous natural product, to quite different extents. Ultimately, an optimized constructed strain containing 15 exogenous genes achieved the highest yield of 144.58 mg/g DCW and produced a lycopene titer of 2144.83 mg/L in a 5 L bioreactor. Our strategy significantly expands the capability of Y. lipolytica genetic manipulation and metabolic engineering.
期刊介绍:
The journal is particularly interested in studies on the design and synthesis of new genetic circuits and gene products; computational methods in the design of systems; and integrative applied approaches to understanding disease and metabolism.
Topics may include, but are not limited to:
Design and optimization of genetic systems
Genetic circuit design and their principles for their organization into programs
Computational methods to aid the design of genetic systems
Experimental methods to quantify genetic parts, circuits, and metabolic fluxes
Genetic parts libraries: their creation, analysis, and ontological representation
Protein engineering including computational design
Metabolic engineering and cellular manufacturing, including biomass conversion
Natural product access, engineering, and production
Creative and innovative applications of cellular programming
Medical applications, tissue engineering, and the programming of therapeutic cells
Minimal cell design and construction
Genomics and genome replacement strategies
Viral engineering
Automated and robotic assembly platforms for synthetic biology
DNA synthesis methodologies
Metagenomics and synthetic metagenomic analysis
Bioinformatics applied to gene discovery, chemoinformatics, and pathway construction
Gene optimization
Methods for genome-scale measurements of transcription and metabolomics
Systems biology and methods to integrate multiple data sources
in vitro and cell-free synthetic biology and molecular programming
Nucleic acid engineering.