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.
- xylose,
- zeaxanthin,
- DNA,
- biosynthesis,
- chemical analysis,
- chromosome,
- functional genomics,
- fungal strain,
- homologous recombination,
- metabolic engineering,
- molecular cloning,
- gene vector,
- genetics,
- Institute for Genomic Biology
Available at: http://works.bepress.com/zengyi_shao/2/