Precise Manipulation of Chromosomes in Vivo Enables Genome-Wide Codon Replacement

• DOI: 10.1126/science.1205822
• Publication Date: 2011-07-14T18:53:28Z
• Authors: Farren J. Isaacs; Peter A. Carr; Harris H. Wang; Marc J. Lajoie; Bram Sterling; Laurens Kraal; Andrew C. Tolonen; Tara A. Gianoulis; Daniel B. Goodman; Nikos B. Reppas; Christopher J. Emig; Duhee Bang; Samuel J. Hwang; Michael C. Jewett; Joseph M. Jacobson; George M. Church
• Keywords: Genome Engineering; Stop codon

We present genome engineering technologies that are capable of fundamentally reengineering genomes from the nucleotide to megabase scale. used multiplex automated (MAGE) site-specifically replace all 314 TAG stop codons with synonymous TAA in parallel across 32 Escherichia coli strains. This approach allowed us measure individual recombination frequencies, confirm viability for each modification, and identify associated phenotypes. developed hierarchical conjugative assembly (CAGE) merge these sets codon modifications into 80 precise changes, which demonstrate substitutions can be combined higher-order strains without synthetic lethal effects. Our methods treat chromosome as both an editable evolvable template, permitting exploration vast genetic landscapes.