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• W2890901605 abstract "This work is divided into two parts. The first examines the role of Wnt signaling and the Tcf/Lef transcription factors in determining mouse embryonic stem cell (mESC) fate decisions. I found that Tcf/Lef factors are not required for the maintenance of the naive mESC state. In contrast, one of the Tcf/Lef factors, Tcf7l1 (formerly Tcf3), is required for the first step of naive embryonic stem cell differentiation. The mechanism by which Wnt signaling inhibits this transition through inactivation of Tcf7l1 is delineated first in mESC. In addition to mESC, examples of Tcf7l1 inactivation by Wnt signaling are presented in aggressive breast malignancies and throughout embryonic development, such as in eyelid closure and the developing forebrain, suggesting widespread applicability. The second part of this work examines methods for improving the efficiency of generating novel genetically modified mESC lines. I make use of CRISPR/Cas9 based tools to introduce double stranded breaks in genomic DNA with subsequent stimulation of homology directed repair (HDR). The basic characteristics of HDR donor DNAs were first identified and optimized, including homology arm length, insert size, and concentration of DNA. In addition, I characterized a basic property of heterogeneous cell populations, whereby a cell that undergoes one successful HDR event is much more likely to perform a second. This phenomenon was labeled co-incident insertion (COIN). Finally, I developed a series of constructs that make use COIN to increase the frequency of desired mutations and streamline the genome editing process." @default.
• W2890901605 created "2018-09-27" @default.
• W2890901605 creator A5054844413 @default.
• W2890901605 date "2017-10-27" @default.
• W2890901605 modified "2023-09-26" @default.
• W2890901605 title "Genome Editing and Differentiation of Embryonic Stem Cells" @default.
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