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. 2015 Jun 18;43(13):6359–6372. doi: 10.1093/nar/gkv621

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© The Author(s) 2015. Published by Oxford University Press on behalf of Nucleic Acids Research.

This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/4.0/), which permits unrestricted reuse, distribution, and reproduction in any medium, provided the original work is properly cited.

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Figure 5. — The fidelity of end-joining in SMARCAL1 mutant cells. (A) Schematic diagram showing DSB-repair events that repair I-Sce1-induced DSBs in the endogenous thymidine kinase (TK) locus. TK^+/− cells carry an I-Sce1 site in intronic sequences of the wild-type TK allele and a mutation in exon4 of the mutant TK allele. DSB repair associated with deletion in exon5 coding sequences would yield TK^−/− clones from TK^+/− cells. The number of TK^−/− clones was measured by counting the number of trifluorothymidine (TFT)-resistant colonies. (B) Histogram representing the frequency of DSB-repair events (y-axis) in the indicated genotypes (x-axis). Error bars indicate SD of more than three independent experiments. P-value was calculated by Student's t-test. (C) Box plot representing the length of nucleotide deletion (y-axis) in the indicated genotypes (x-axis). PCR was performed from genomic DNA isolated from at least 50 TFT-resistant clones of each genotype, as shown in Supplementary Figure S6. (D) Schematic representation of the structure of wild-type, R764Q and Δ30 Smarcal1 proteins. SMARCAL1^−/− TK6 cells were reconstituted with SMARCAL1^WT, SMARCAL1^Δ30 or SMARCAL1^R764Q transgene. Western blot analysis for the expression of individual transgenes in SMARCAL1^−/− cells. β-actin was used as a loading control. (E) Histogram representing the frequency of TFT-resistant colonies (y-axis) in the indicated genotypes (x-axis). Error bars indicate SD of more than three independent experiments.