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Updated: Jun 27, 2026

Rapid Characterization of Genetic Parts with Cell-Free Systems
Published on: August 30, 2021
DNA assembler, an in vivo genetic method for rapid construction of biochemical pathways
Zengyi Shao1, Hua Zhao, Huimin Zhao
1Department of Chemical and Biomolecular Engineering, Institute for Genomic Biology, University of Illinois at Urbana-Champaign, Urbana, IL 61801, USA.
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 (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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