title: Type IIs restriction based combinatory modulation technique for metabolic pathway optimization.
abstract: BACKGROUND
  One of the most important research subjects of metabolic engineering is pursuing a balanced metabolic pathway, which is the basis of an efficient cell factory. In this work, we dedicated to develop a simple and efficient technique to modulate expression of multiple genes simultaneously, and select for the optimal regulation pattern.
  
  
  RESULTS
  A Type IIs restriction based combinatory modulation (TRCM) technique was designed and established in the research. With this technique, a plasmid library containing variably regulated mvaE, mvaS, mvaK1, mvaD and mvaK2of the mevalonate (MVA) pathway were obtained and transformed into E. coli DXS37-IDI46 to obtain a β-carotene producer library. The ratio of successfully assembled plasmids was determined to be 35%, which was increased to 100% when color based pre-screening was applied. Representative strains were sequenced to contain diverse RBSs as designed to regulate expression of MVA pathway genes. A relatively balanced MVA pathway was achieved in E. coli cell factory to increase the β-carotene yield by two fold. Furthermore, the approximate regulation pattern of this optimal MVA pathway was illustrated.
  
  
  CONCLUSIONS
  A TRCM technique for metabolic pathway optimization was designed and established in this research, which can be applied to various applications in terms of metabolic pathway regulation and optimization.
authors.affiliation: Tianjin Institute of Industrial Biotechnology, Chinese Academy of Sciences, Tianjin, 300308, People's Republic of China.|Key Laboratory of Systems Microbial Biotechnology, Chinese Academy of Sciences, Tianjin, 300308, People's Republic of China.
authors.forename: Lijun
authors.initials: L
authors.lastname: Ye
authors.affiliation: Tianjin Institute of Industrial Biotechnology, Chinese Academy of Sciences, Tianjin, 300308, People's Republic of China.|Key Laboratory of Systems Microbial Biotechnology, Chinese Academy of Sciences, Tianjin, 300308, People's Republic of China.|School of Pharmacy, East China University of Science and Technology, Shanghai, 200237, People's Republic of China.
authors.forename: Ping
authors.initials: P
authors.lastname: He
authors.affiliation: Tianjin Institute of Industrial Biotechnology, Chinese Academy of Sciences, Tianjin, 300308, People's Republic of China.|Key Laboratory of Systems Microbial Biotechnology, Chinese Academy of Sciences, Tianjin, 300308, People's Republic of China.
authors.forename: Qingyan
authors.initials: Q
authors.lastname: Li
authors.affiliation: Tianjin Institute of Industrial Biotechnology, Chinese Academy of Sciences, Tianjin, 300308, People's Republic of China. bi_ch@tib.cas.cn.|Key Laboratory of Systems Microbial Biotechnology, Chinese Academy of Sciences, Tianjin, 300308, People's Republic of China. bi_ch@tib.cas.cn.
authors.forename: Xueli
authors.initials: X
authors.lastname: Zhang
authors.affiliation: Tianjin Institute of Industrial Biotechnology, Chinese Academy of Sciences, Tianjin, 300308, People's Republic of China. zhang_xl@tib.cas.cn.|Key Laboratory of Systems Microbial Biotechnology, Chinese Academy of Sciences, Tianjin, 300308, People's Republic of China. zhang_xl@tib.cas.cn.
authors.forename: Changhao
authors.identifier: http://orcid.org/0000-0002-1940-8511
authors.initials: C
authors.lastname: Bi
mesh_terms: D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D015723:Gene Library; D060847:Metabolic Engineering / Q000379:methods*; D053858:Metabolic Networks and Pathways / Q000235:genetics; D008798:Mevalonic Acid / Q000378:metabolism; D010957:Plasmids; D019207:beta Carotene / Q000096:biosynthesis
chemical_list: D019207:beta Carotene; D008798:Mevalonic Acid
keywords: MVA pathway; Metabolic pathway optimization; Terpene; Type IIs restriction; β-carotene