title: Yeast Pathway Kit: A Method for Metabolic Pathway Assembly with Automatically Simulated Executable Documentation.
abstract: We have developed the Yeast Pathway Kit (YPK) for rational and random metabolic pathway assembly in Saccharomyces cerevisiae using reusable and redistributable genetic elements. Genetic elements are cloned in a suicide vector in a rapid process that omits PCR product purification. Single-gene expression cassettes are assembled in vivo using genetic elements that are both promoters and terminators (TP). Cassettes sharing genetic elements are assembled by recombination into multigene pathways. A wide selection of prefabricated TP elements makes assembly both rapid and inexpensive. An innovative software tool automatically produces detailed self-contained executable documentation in the form of pydna code in the narrative Jupyter notebook format to facilitate planning and sharing YPK projects. A d-xylose catabolic pathway was created using YPK with four or eight genes that resulted in one of the highest growth rates reported on d-xylose (0.18 h(-1)) for recombinant S. cerevisiae without adaptation. The two-step assembly of single-gene expression cassettes into multigene pathways may improve the yield of correct pathways at the cost of adding overall complexity, which is offset by the supplied software tool.
authors.affiliation: CBMA-Centre of Molecular and Environmental Biology, Department of Biology, University of Minho , Campus de Gualtar, Braga 4710-057, Portugal.
authors.forename: Filipa
authors.initials: F
authors.lastname: Pereira
authors.affiliation: CBMA-Centre of Molecular and Environmental Biology, Department of Biology, University of Minho , Campus de Gualtar, Braga 4710-057, Portugal.
authors.forename: Flávio
authors.initials: F
authors.lastname: Azevedo
authors.affiliation: Division of Applied Microbiology, Department of Chemistry, Lund University , SE-22100 Lund, Sweden.
authors.forename: Nadia Skorupa
authors.initials: NS
authors.lastname: Parachin
authors.affiliation: Division of Applied Microbiology, Department of Chemistry, Lund University , SE-22100 Lund, Sweden.
authors.forename: Bärbel
authors.initials: B
authors.lastname: Hahn-Hägerdal
authors.affiliation: Division of Applied Microbiology, Department of Chemistry, Lund University , SE-22100 Lund, Sweden.
authors.forename: Marie F
authors.initials: MF
authors.lastname: Gorwa-Grauslund
authors.affiliation: CBMA-Centre of Molecular and Environmental Biology, Department of Biology, University of Minho , Campus de Gualtar, Braga 4710-057, Portugal.
authors.forename: Björn
authors.initials: B
authors.lastname: Johansson
mesh_terms: D004282:Documentation / Q000379:methods; D015870:Gene Expression / Q000235:genetics; D053858:Metabolic Networks and Pathways / Q000235:genetics*; D011401:Promoter Regions, Genetic / Q000235:genetics; D012441:Saccharomyces cerevisiae / Q000235:genetics*; D013728:Terminator Regions, Genetic / Q000235:genetics; D014994:Xylose / Q000235:genetics
chemical_list: D014994:Xylose
keywords: Saccharomyces cerevisiae; bioinformatics; d-xylose; metabolic engineering; synthetic biology