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. 2017 Oct 20;45(20):11837–11857. doi: 10.1093/nar/gkx847

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© The Author(s) 2017. 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-nc/4.0/), which permits non-commercial re-use, distribution, and reproduction in any medium, provided the original work is properly cited. For commercial re-use, please contact journals.permissions@oup.com

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Figure 6. — FANCD2 plays a more crucial role than FANCI to promote replication fork recovery. (A) Schematic of the replication fork restart protocol with representative images of DNA fibers. Red tracks: DigU; green tracks: BioU. (B) FANCD2 plays a more crucial role than FANCI in promoting the restart of APH-stalled replication forks. The efficiency of replication restart in WT, D2^−/− (clone #39), I^−/− (clone #28), and ID2 DKO (clone #1), as well as in the complemented cells was measured as the number of restarted replication forks after APH (30 μM) mediated fork stalling (DigU-BioU tracts), compared with the total number of DigU-labeled tracts (DigU plus DigU-BioU). Statistical significance at P < 0.05, P < 0.01 and P < 0.001 are indicated as *, **, ***, respectively. (C) FANCD2 plays a more crucial role than FANCI in suppressing new origin firing in the presence of stalled replication forks. WT, D2^−/− (clone #39), I^−/− (clone #28), and ID2 DKO (clone #1), as well as the complemented cells were analyzed for the fraction of newly fired replication origins during the 40 min post-APH recovery period. Fractions were measured as the number of green-only (BioU) tracts compared with the total number of spreading replication tracts (BioU plus DigU-BioU). Statistical significance at P < 0.05, P < 0.01 and P < 0.001 are indicated as *, **, ***, respectively. (D) Schematic of the DNA combing assay with representative images of stretched DNA fibers. Green tracks: CldU; red tracks: IdU. (E) FANCD2 plays a more crucial role than FANCI in promoting the restart of HU-stalled replication forks. The efficiency of replication restart in WT, D2^−/−(clones #29 and #39), I^−/− (clones #28 and #30) and ID2 DKO (clones #1 and #2) was measured as the number of restarted replication forks after HU-mediated fork stalling (CldU-IdU tracts), compared with the total number of CldU-labeled tracts (CldU plus CldU-IdU). Data points were averaged between clones of identical genetic backgrounds. Statistical significance at P < 0.05, P < 0.01 and P < 0.001 are indicated as *, **, ***, respectively.