Enhanced multiplex genome engineering through co-operative oligonucleotide co-selection.
Genome-scale engineering of living organisms requires precise and economical methods to efficiently modify many loci within chromosomes. One such example is the directed integration of chemically synthesized single-stranded deoxyribonucleic acid (oligonucleotides) into the chromosome of Escherichia coli during replication. Herein, we present a general co-selection strategy in multiplex genome engineering that yields highly modified cells. We demonstrate that disparate sites throughout the genome can be easily modified simultaneously by leveraging selectable markers within 500 kb of the target sites. We apply this technique to the modification of 80 sites in the E. coli genome.
The Center for Bits and Atoms, Massachusetts Institute of Technology, Cambridge, MA 02139, USA. carr@ll.mit.edu
Peter A
PA
Carr
Harris H
HH
Wang
Bram
B
Sterling
Farren J
FJ
Isaacs
Marc J
MJ
Lajoie
George
G
Xu
George M
GM
Church
Joseph M
JM
Jacobson
D002876:Chromosomes, Bacterial; D004926:Escherichia coli / Q000235:genetics; D005818:Genetic Engineering / Q000379:methods*; D016680:Genome, Bacterial; D009841:Oligonucleotides / Q000737:chemistry*
D009841:Oligonucleotides