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. 2019 Oct 1;38(24):e101751. doi: 10.15252/embj.2019101751

Figure 2. Effect of the distance between the DSBs and the repeats on RMD frequency.

Figure 2

  • A
    A diagram indicating the position of DSBs generated by sgRNAs/Cas9 on the EGFP‐SSA reporter.
  • B
    RMD efficiency is decreased when the distance from a DSB to one repeat is increased. RMD assays were performed in U2OS (EGFP‐SSA‐3427‐17) cells using the indicated sgRNAs/Cas9. The distance between the sgRNA/Cas9 cleavage sites and the repeat sequences is shown.
  • C, D
    Knockdown of POLD3 impairs RMD when a DSB is close to one repeat. RMD frequency was determined in U2OS (EGFP‐SSA‐3427‐17) cells using the indicated sgRNAs/Cas9 with or without depletion of POLD3 by lentivirus‐expressed shRNA. Cells infected with a lentiviral empty vector were used as control. The distance between the sgRNA/Cas9 cleavage sites and the repeat sequences is indicated. The knockdown level of POLD3 was shown by Western blot, and KU70 was used as the loading control.
Data information: Error bars represent standard deviation (SD) of at least three independent experiments. *P < 0.05, **P < 0.01, ns, not significant, two‐tailed non‐paired t‐test.