DNA assembler, an in vivo genetic method for rapid construction of biochemical pathways.
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.
Shao
Zengyi
Z
Department of Chemical and Biomolecular Engineering, Institute for Genomic Biology, University of Illinois at Urbana-Champaign, Urbana, IL 61801, USA.
Zhao
Hua
H
Zhao
Huimin
H
D053898:Biosynthetic Pathways / Q000235:genetics; D003001:Cloning, Molecular / Q000379:methods*; D004247:DNA / Q000378:metabolism; D005800:Genes, Fungal; D005822:Genetic Vectors; D053858:Metabolic Networks and Pathways / Q000235:genetics*; D010957:Plasmids / Q000235:genetics; D016133:Polymerase Chain Reaction; D011995:Recombination, Genetic; D012441:Saccharomyces cerevisiae / Q000235:genetics* / Q000378:metabolism; D024341:Xanthophylls / Q000096:biosynthesis; D014994:Xylose / Q000378:metabolism; D065146:Zeaxanthins
D024341:Xanthophylls; D065146:Zeaxanthins; D004247:DNA; D014994:Xylose