Scharenberg, Andrew MGwiazda, Kamila2014-10-132014-10-132014-10-132014Gwiazda_washington_0250E_13247.pdfhttp://hdl.handle.net/1773/26386Thesis (Ph.D.)--University of Washington, 2014Targeted genome engineering is a technology that has the potential to vastly improve clinical medicine, as well as biological research. With the ability to design enzymes targeting any given DNA locus of interest, and mutate, delete, or repair the sequence, we can manipulate the genome to a degree that was not possible even a decade ago. However, both the rate at which these desired modifications occur following a nuclease-induced double strand break, as well as the type of repair that occurs, are variables we do not yet have full control over. In this work, we have found that introducing exogenous DNA end-modifying enzymes such as Trex2 and Artemis has a significant impact on promoting one type of repair, NHEJ. In contrast, altering the structure of the donor DNA by linearizing it in vivo promotes repair by HR. Additionally, we present evidence that not all designer endonuclease breaks behave similarly: those that leave 3' overhangs are much more amenable to repair by HR and SSA than those with 5' overhangs.application/pdfen-USCopyright is held by the individual authors.Genome engineeringMolecular biologyImmunologymolecular and cellular biologyThesis