title: YeastFab: the design and construction of standard biological parts for metabolic engineering in Saccharomyces cerevisiae.
abstract: It is a routine task in metabolic engineering to introduce multicomponent pathways into a heterologous host for production of metabolites. However, this process sometimes may take weeks to months due to the lack of standardized genetic tools. Here, we present a method for the design and construction of biological parts based on the native genes and regulatory elements in Saccharomyces cerevisiae. We have developed highly efficient protocols (termed YeastFab Assembly) to synthesize these genetic elements as standardized biological parts, which can be used to assemble transcriptional units in a single-tube reaction. In addition, standardized characterization assays are developed using reporter constructs to calibrate the function of promoters. Furthermore, the assembled transcription units can be either assayed individually or applied to construct multi-gene metabolic pathways, which targets a genomic locus or a receiving plasmid effectively, through a simple in vitro reaction. Finally, using β-carotene biosynthesis pathway as an example, we demonstrate that our method allows us not only to construct and test a metabolic pathway in several days, but also to optimize the production through combinatorial assembly of a pathway using hundreds of regulatory biological parts.
authors.affiliation: MOE Key Laboratory of Bioinformatics, School of Life Sciences, Tsinghua University, Beijing 100084, China.
authors.forename: Yakun
authors.initials: Y
authors.lastname: Guo
authors.affiliation: MOE Key Laboratory of Bioinformatics, School of Life Sciences, Tsinghua University, Beijing 100084, China.
authors.forename: Junkai
authors.initials: J
authors.lastname: Dong
authors.affiliation: MOE Key Laboratory of Bioinformatics, School of Life Sciences, Tsinghua University, Beijing 100084, China.
authors.forename: Tong
authors.initials: T
authors.lastname: Zhou
authors.affiliation: School of Biological Sciences, The King's Buildings, University of Edinburgh, Edinburgh EH9 3BF, United Kingdom.
authors.forename: Jamie
authors.initials: J
authors.lastname: Auxillos
authors.affiliation: MOE Key Laboratory of Bioinformatics, School of Life Sciences, Tsinghua University, Beijing 100084, China.
authors.forename: Tianyi
authors.initials: T
authors.lastname: Li
authors.affiliation: MOE Key Laboratory of Bioinformatics, School of Life Sciences, Tsinghua University, Beijing 100084, China.
authors.forename: Weimin
authors.initials: W
authors.lastname: Zhang
authors.affiliation: MOE Key Laboratory of Bioinformatics, School of Life Sciences, Tsinghua University, Beijing 100084, China.
authors.forename: Lihui
authors.initials: L
authors.lastname: Wang
authors.affiliation: School of Biological Sciences, The King's Buildings, University of Edinburgh, Edinburgh EH9 3BF, United Kingdom.
authors.forename: Yue
authors.initials: Y
authors.lastname: Shen
authors.affiliation: School of Biological Sciences, The King's Buildings, University of Edinburgh, Edinburgh EH9 3BF, United Kingdom.
authors.forename: Yisha
authors.initials: Y
authors.lastname: Luo
authors.affiliation: School of Biological Sciences, The King's Buildings, University of Edinburgh, Edinburgh EH9 3BF, United Kingdom.
authors.forename: Yijing
authors.initials: Y
authors.lastname: Zheng
authors.affiliation: Wuxi Qinglan Biotechnology Inc., Yixing, Jiangsu 214200, China.
authors.forename: Jiwei
authors.initials: J
authors.lastname: Lin
authors.affiliation: MOE Key Laboratory of Bioinformatics, School of Life Sciences, Tsinghua University, Beijing 100084, China.
authors.forename: Guo-Qiang
authors.initials: GQ
authors.lastname: Chen
authors.affiliation: MOE Key Laboratory of Bioinformatics, School of Life Sciences, Tsinghua University, Beijing 100084, China.
authors.forename: Qingyu
authors.initials: Q
authors.lastname: Wu
authors.affiliation: School of Biological Sciences, The King's Buildings, University of Edinburgh, Edinburgh EH9 3BF, United Kingdom yizhi.cai@ed.ac.uk.
authors.forename: Yizhi
authors.initials: Y
authors.lastname: Cai
authors.affiliation: MOE Key Laboratory of Bioinformatics, School of Life Sciences, Tsinghua University, Beijing 100084, China jbdai@biomed.tsinghua.edu.cn.
authors.forename: Junbiao
authors.initials: J
authors.lastname: Dai
mesh_terms: D060847:Metabolic Engineering / Q000379:methods*; D053858:Metabolic Networks and Pathways / Q000235:genetics*; D012441:Saccharomyces cerevisiae / Q000235:genetics* / Q000378:metabolism; D014158:Transcription, Genetic; D019207:beta Carotene / Q000096:biosynthesis
chemical_list: D019207:beta Carotene