Research Paper Volume 2, Issue 9 pp 582—596

ATM-independent, high-fidelity nonhomologous end joining predominates in human embryonic stem cells

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Figure 3. NHEJ occurs with faster kinetics after terminal differentiation. (A) hESCs, NPs and astrocytes were seeded and 12 h later infected with Ad-SceI at an MOI of 100. Expression of HA-tagged I-SceI was examined in samples harvested 24 h after infection. (B) BG01V/-, NP/-, and astrocyte/NHEJ-red cells were infected with Ad-SceI and collected 24 h later. (Columns) Relative NHEJ levels were determined by genomic DNA qPCR and normalized to β-actin levels; (Error bars) SEM for data sets n = 3. Fold (x) indicates changes in the relative repair levels when compared to the hESC sample. *p < 0.05: **p < 0.01; ***p < 0.001. (C) (Top Panel) BG01V/-, NP/-, and astrocyte/NHEJ-red cells were infected with Ad-I-SceI at an MOI of 30. DsRed events were determined by FACS 48 and 72 h after infection. Fold (x) and statistical significance indicates changes in the relative repair levels when compared to the hESC sample. (Columns) % DsRed+ cells with 60,000 events collected; (Error bars) SEM for three independent experiments. (Bottom Panel) Representative FACS images of DsRed+ cells at 72 h after infection.