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Nucleic Acids Res. 2009 Feb;37(2):e16. doi: 10.1093/nar/gkn991. Epub 2008 Dec 12.

DNA assembler, an in vivo genetic method for rapid construction of biochemical pathways.

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Department of Chemical and Biomolecular Engineering, Institute for Genomic Biology, University of Illinois at Urbana-Champaign, Urbana, IL 61801, USA.


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 (approximately 9 kb DNA consisting of three genes), a functional zeaxanthin biosynthesis pathway (approximately 11 kb DNA consisting of five genes) and a functional combined D-xylose utilization and zeaxanthin biosynthesis pathway (approximately 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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