DNA assembler,an in vivo genetic method for rapid construction of biochemical pathways |
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| Authors: | Zengyi Shao Hua Zhao Huimin Zhao |
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| Institution: | 1.Department of Chemical and Biomolecular Engineering and 2.Department of Chemistry, Biochemistry, and Bioengineering, Institute for Genomic Biology, University of Illinois at Urbana-Champaign, Urbana, IL 61801, USA |
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| Abstract: | The assembly of large recombinant DNA encoding a whole biochemical pathway or genome represents a significant challenge. Here, we report a new method, DNA assembler, which allows the assembly of an entire biochemical pathway in a single step via in vivo homologous recombination in Saccharomyces cerevisiae. We show that DNA assembler can rapidly assemble a functional d-xylose utilization pathway (∼9 kb DNA consisting of three genes), a functional zeaxanthin biosynthesis pathway (∼11 kb DNA consisting of five genes) and a functional combined d-xylose utilization and zeaxanthin biosynthesis pathway (∼19 kb consisting of eight genes) with high efficiencies (70–100%) either on a plasmid or on a yeast chromosome. As this new method only requires simple DNA preparation and one-step yeast transformation, it represents a powerful tool in the construction of biochemical pathways for synthetic biology, metabolic engineering and functional genomics studies. |
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