PubAg

Main content area

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

Author:
Shao, Zengyi, Zhao, Hua, Zhao, Huimin
Source:
Nucleic acids research 2009 v.37 no.2 pp. e16
ISSN:
0305-1048
Subject:
Saccharomyces cerevisiae, biochemical pathways, biosynthesis, chromosomes, genes, genomics, homologous recombination, metabolic engineering, methodology, plasmids, recombinant DNA, synthetic biology, xylose, yeasts, zeaxanthin
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.
Agid:
2122920