Chapter 2: Trackable Multiplex Recombineering (TRMR) and Next-Generation Genome Design Technologies: Modifying Gene Expression in E. coli by Inserting Synthetic DNA Cassettes and Molecular Barcodes

Emily Freed, Carrie Eckert, Gur Pines, Ryan Gill

Research output: Chapter in Book/Report/Conference proceedingChapter

Abstract

This chapter describes the trackable multiplex recombineering (TRMR) and tunable trackable multiplex recombineering (T2RMR) techniques, which not only make the multiplexing of recombineering possible in Escherichia coli but also provide the ability to track the engineered genetic changes accurately. TRMR and T2RMR have only been used for modifying the expression level of genes. Screening and selection of TRMR and T2RMR clones can be carried out using either liquid or solid media with any chemical compound that modifies growth or confers a phenotype that can be detected by a high-throughput assay. In order to engineer a genome using TRMR or T2RMR, a synthetic DNA (synDNA) cassette is created that encodes for a genetic feature and a molecular barcode that is used to track each feature. These synDNA cassettes are then introduced in parallel into cell populations via recombineering.
Original languageAmerican English
Title of host publicationSynthetic Biology: Parts, Devices and Applications
EditorsC. Smolke
Pages15-31
DOIs
StatePublished - 2018

NREL Publication Number

  • NREL/CH-2700-72219

Keywords

  • E. coli
  • gene expression
  • genetic feature
  • molecular barcodes
  • next-generation genome design technologies
  • synthetic DNA cassette
  • tunable trackable multiplex recombineering

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