Construction and engineering of large biochemical pathways via DNA assembler

Zengyi Shao, Huimin Zhao

Research output: Chapter in Book/Report/Conference proceedingChapter

Abstract

DNA assembler enables rapid construction and engineering of biochemical pathways in a one-step fashion by exploitation of the in vivo homologous recombination mechanism in Saccharomyces cerevisiae. It has many applications in pathway engineering, metabolic engineering, combinatorial biology, and synthetic biology. Here we use two examples including the zeaxanthin biosynthetic pathway and the aureothin biosynthetic gene cluster to describe the key steps in the construction of pathways containing multiple genes using the DNA assembler approach. Methods for construct design, pathway assembly, pathway confirmation, and functional analysis are shown. The protocol for fine genetic modifications such as site-directed mutagenesis for engineering the aureothin gene cluster is also illustrated.

Original languageEnglish (US)
Title of host publicationSynthetic Biology
EditorsKaren Polizzi, Cleo Kontoravdi
Pages85-106
Number of pages22
DOIs
StatePublished - 2013

Publication series

NameMethods in Molecular Biology
Volume1073
ISSN (Print)1064-3745

Keywords

  • DNA assembler
  • Gene cluster characterization and engineering
  • In vivo homologous recombination
  • Metabolic engineering
  • Pathway engineering
  • Synthetic biology

ASJC Scopus subject areas

  • Molecular Biology
  • Genetics

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  • Cite this

    Shao, Z., & Zhao, H. (2013). Construction and engineering of large biochemical pathways via DNA assembler. In K. Polizzi, & C. Kontoravdi (Eds.), Synthetic Biology (pp. 85-106). (Methods in Molecular Biology; Vol. 1073). https://doi.org/10.1007/978-1-62703-625-2_9