title: Improved Gene Targeting through Cell Cycle Synchronization.
abstract: Gene targeting is a challenge in organisms where non-homologous end-joining is the predominant form of recombination. We show that cell division cycle synchronization can be applied to significantly increase the rate of homologous recombination during transformation. Using hydroxyurea-mediated cell cycle arrest, we obtained improved gene targeting rates in Yarrowia lipolytica, Arxula adeninivorans, Saccharomyces cerevisiae, Kluyveromyces lactis and Pichia pastoris demonstrating the broad applicability of the method. Hydroxyurea treatment enriches for S-phase cells that are active in homologous recombination and enables previously unattainable genomic modifications.
authors.affiliation: Total New Energies, Emeryville, California, United States of America.
authors.forename: Vasiliki
authors.initials: V
authors.lastname: Tsakraklides
authors.affiliation: Total New Energies, Emeryville, California, United States of America.
authors.forename: Elena
authors.initials: E
authors.lastname: Brevnova
authors.affiliation: Novogy Inc., Cambridge, Massachusetts, United States of America; Department of Chemical Engineering, Massachusetts Institute of Technology, Cambridge, Massachusetts, United States of America.
authors.forename: Gregory
authors.initials: G
authors.lastname: Stephanopoulos
authors.affiliation: Novogy Inc., Cambridge, Massachusetts, United States of America.
authors.forename: A Joe
authors.initials: AJ
authors.lastname: Shaw
mesh_terms: D002453:Cell Cycle / Q000235:genetics*; D003001:Cloning, Molecular; D018390:Gene Targeting / Q000379:methods*; D007716:Kluyveromyces / Q000235:genetics; D010843:Pichia / Q000235:genetics; D012441:Saccharomyces cerevisiae / Q000235:genetics; D004718:Saccharomycetales / Q000235:genetics; D025062:Yarrowia / Q000235:genetics